Dissertations / Theses on the topic 'E1 and E2'
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Hudson, Natalia Joanna. "Analysis of diversity of hepatitis C virus glycoproteins E1 and E2." Thesis, University of Nottingham, 2012. http://eprints.nottingham.ac.uk/12644/.
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Hepatitis C Virus (HCV) exists as a population of sequence variants that evolves during infection adapting to host pressures. The main targets for the immune response are the envelope glycoproteins E1 and E2, which also mediate viral cell entry. The first hypervariable region (HVR1) of E2, previously implicated in the outcome of acute infection, has been a focus of many studies. However more broadly neutralising antibodies tend to target epitopes outside this region, yet evolution of full length E1E2 heterodimer is poorly understood. The HCV transmission and window period as well as seroconversion are the evolutionary events shaping primary infection hence influencing outcome of acute infection. However, due to the asymptomatic character of the early phases of HCV infection, evolutionary data describing this interval is still lacking depth. Defining the genetic and phenotypic characteristics of HCV population of sequence variants that establish infection in a new host would aid vaccine and new therapy design. This study aimed to identify patterns of HCV envelope glycoprotein evolution upon transmission and during early phases of disease. We studied this in three settings: experimental transmission of immunocompromised mice by known inoculum; occurrence of horizontal transmission in a haemodialysis unit between hypothesised source and index case individuals; and unrelated cases of acutely infected HCV patients. The single genome amplification (SGA) approach was utilised, which allowed us to accurately assign linkage between substitutions and determine the frequency distribution of E1E2 variants in analysed viral populations. Data from the first experimental setting indicates that a selective sweep occurs upon HCV transmission, with selective amplification of envelope sequence variants that possess fitness advantage at entry level. Molecular determinants associated with this enhanced infectivity have also been identified. In further part of the project we confirmed a horizontal infection in haemodialysis unit with use of phylogenetic methods and suggested revision of current safety guidelines. Analysis of sequences from the last setting showed that indeed HVR1 might not be a good enough indicator of evolutionary events in the acute phase, as linked substitutions occur also outside this region. Seroconversion is associated with increasing population diversity indicating role of antibodies in driving HCV evolution, which is host specific.
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Sorathia, Rina. "Identification of human papillomavirus type 16 (HPV16) E1^E4 binding partners and the characterisation of the E1^E4/E2 interaction." Thesis, University College London (University of London), 2005. http://discovery.ucl.ac.uk/1446481/.
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Human papillomavirus type 16 (HPV16) is a DNA tumour virus that can infect epithelial tissue and can cause hyperproliferative lesions that can progress to cancer. The expression of the HPV late protein, E1.;E4 is lost during malignantprogression, but in productive infections E1.;E4 is abundantly expressed inthe late stages of HPV infection. The appearance of E1.;E4 coincides with theonset of genome amplification and the upregulation of the viral replication proteins El and E2 and precedes the synthesis of the viral capsid proteins L1 and L2. In this study we show that HPV16 E1.;E4 (16E1.;E4) can interact directlywith several cellular and viral proteins including the late proteins L1 and L2 and the early proteins E7 and E2. HPV16 E2 associates with the phosphorylated, unphosphorylated and cleaved forms of 16E1.;E4, with stable association beingdependent on sequences in the central highly charged region and the extreme C-terminus of E1.;E4. We observed that co-expression of E2 and E1.;E4 in culturedepithelial cells resulted in the progressive accumulation of E2 to E1.;E4 boundstructures in the cytoplasm, and a reduction in nuclear E2 levels at late time points. This phenomenon in cells that were co-expressing E2, E1.;E4 andEl, at ratios mimicking those seen in the late stages of the virus life cycle, coincided with an increase in HPV origin-dependent replication and enhanced El/E2-mediated transcription from the wild-type HPV16 early promoter (p97). The co-expression of E1.;E4 with El and E2 at late time points was also shownto reduce p97 activity when E2 was expressed at levels that were optimised for E1/E2- mediated transactivation. We suggest that during productive infection, the ability of E1.;E4 to bind to and alter the distribution of E2 in the cellmay facilitate genome amplification and the expression of S-phase promoting HPV proteins, E6 and E7.
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Cocquerel, Laurence. "Caracterisation des domaines transmembranaires des glycoproteines e1 et e2 du virus de l'hepatite c." Paris 6, 2001. http://www.theses.fr/2001PA066052.
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Dans ce travail de these, nous nous sommes interesses aux etapes precoces de la formation de l'enveloppe virale du virus de l'hepatite c (vhc). Le genome du vhc code deux glycoproteines d'enveloppe appelees e1 et e2. Ces glycoproteines possedent un large ectodomaine n-terminal hautement glycosyle et sont ancrees dans les membranes par leur domaine transmembranaire (tm) c-terminal. Ces deux glycoproteines interagissent de maniere non covalente pour former un heterodimere e1e2 qui est suppose etre le composant proteique majeur de l'enveloppe du vhc. Lorsqu'il est exprime par des vecteurs d'expression heterologue, l'heterodimere e1e2 est retenu au niveau du reticulum endoplasmique (re). Dans notre travail, nous avons dans un premier temps cherche a identifier les determinants impliques dans cette localisation subcellulaire du complexe e1e2. Par le biais de constructions de proteines chimeriques entre les proteines d'enveloppe du vhc et des proteines normalement exportees a la membrane plasmique (cd4 et/ou cd8), nous avons montre que les domaines tms de e1 et e2 sont des signaux de retention stricte dans le re. Par une approche de mutagenese dirigee, nous avons ensuite montre que la presence d'un ou de deux residus charges, localises au milieu des domaines tms de e1 et e2, jouent un role essentiel dans la localisation subcellulaire de ces proteines. De plus, nous avons constate que ces residus charges sont egalement importants pour l'assemblage du complexe e1e2 ainsi que pour la fonction sequence signal situee dans la moitie c-terminale de ces domaines. Enfin, nous avons determine la topologie de ces domaines tms avant et apres clivage de la polyproteine du vhc et nous avons mis en evidence une dynamique structurale dans les etapes initiales de la biogenese des proteines d'enveloppe du vhc. L'ensemble de ce travail nous permet de proposer un modele reliant la multifonctionnalite des domaines tms des proteines d'enveloppe du vhc a leur dynamique structurale.
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Almeida, Ana Isabel Matos de. "Produção de partículas semelhantes a retrovírus como candidatas a vacina para a Hepatite C." Master's thesis, Faculdade de Ciências e Tecnologia, 2012. http://hdl.handle.net/10362/9297.
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Dissertação para obtenção do Grau de Mestre em
Genética Molecular e BiomedicinaVírus da Hepatite C (HCV) infeta mais de 170 milhões de pessoas em todo o mundo, sendo uma das principais causas de cancro do fígado. Como tal, é extremamente importante e desejável a criação de uma vacina. Recentemente têm vindo a ser desenvolvidas e comercializadas vacinas baseadas em partículas semelhantes a vírus (VLPs) capazes de estimular respostas humorais e celulares eficientes. Os objetivos deste trabalho consistiram no melhoramento de linhas celulares produtoras de VLPs derivadas de retrovírus e no melhoramento da qualidade das partículas virais, com o intuito de produzir VLPs candidatas a vacina para a hepatite C. Para tal, utilizaram-se duas metodologias. Um sistema de troca de cassette (RMCE), que é um sistema versátil e rápido para o desenvolvimento de linhas celulares, permitindo obter uma elevada expressão de um gene de interesse, tendo sido utilizado para expressar os epítopos do HCV. Testaram-se dois sistemas, um mediado pela enzima Cre e outro mediado pela enzima Flipase. Para o melhoramento da qualidade das partículas utilizou-se a metodologia de silenciamento por RNAi para eliminar a incorporação da tetraspanina CD63, uma vez que a presença desta proteína nas VLPs poderá desencadear respostas imunológicas inespecíficas. Verificou-se que o sistema de troca de cassette mediado pela enzima Cre não foi eficiente nas condições experimentais utilizadas, não tendo sido possível produzir VLPs pseudotipadas com os epítopos do HCV. No sistema mediado pela enzima Flipase a troca de cassette foi eficiente obtendo-se clones que expressam as proteínas E1 e E2, para a produção de VLPs pseudotipadas com o envelope do vírus HCV. Em relação ao silenciamento da tetraspanina CD63 apesar de serem testadas diversas condições, a percentagem de silenciamento obtida foi muito reduzida e instável, concluindo-se não ser possível realizar o silenciamento desta proteína através das sequências utilizadas.
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Patel, Janisha. "An investigation of the complexes formed between the hepatitis C virus E1 and E2 glycoproteins." Thesis, University of Glasgow, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.342004.
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Meunier, Jean-Christophe. "Étude des glycoprotéines E1 et E2 du virus de l'hépatite C : influence de la glycosylation de la protéine E1 sur la formation du complexe E1E2." Lille 1, 1999. https://pepite-depot.univ-lille.fr/LIBRE/Th_Num/1999/50376-1999-23.pdf.
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L'infection par le virus de l'hépatite C représente un problème majeur de santé publique. La prévalence dans la population mondiale serait de 0,5% à 1%. L'infection peut se manifester par une hépatite aiguë, une hépatite chronique qui peut dégénérer en cirrhose ou en carcinome hépatocellulaire (plus de 5% des cas). Bien que l'agent infectieux soit cloné depuis 1989, il est actuellement difficile de le propager en culture cellulaire. Par analogie avec la plupart des protéines exposées à la surface des virus, les glycoprotéines d'enveloppe E1 et E2 jouent probablement un rôle important dans l'infection virale en étant responsable de l'interaction du virus avec son ou ses récepteurs à la surface des cellules cibles, ces protéines sont également importantes pour l'immunogénicité virale. L'étude de la maturation et de l'expression des protéines E1 et E2, tout comme leur interaction et leur assemblage est donc nécessaire pour une meilleure compréhension du cycle viral et de la pathogenèse du VHC. Dans un premier temps, les mécanismes de la rétention des protéines E1 et E2, ainsi que du complexe E1E2 à l'intérieur du réticulum endoplasmique (RE) ont été explorés. Il a été montré que le complexe forme par les glycoprotéines E1 et E2 comprend des signaux de rétention dans le RE, portés par les domaines transmembranaires respectifs de ces deux protéines. Ces domaines transmembranaires sont situés respectivement pour les protéines E1 et E2, entre les acides aminés 353 et 384 pour l'un et 718 et 747 pour l'autreEn second lieu, nous avons identifié les sites potentiels de glycosylation reconnus sur la glycoprotéine E1 du virus de l'hépatite C. Nous avons montré que seul le site de glycosylation n°5 de la protéine E1 n'est pas utilisé pour l'addition d'un oligosaccharide. Puis nous avons étudié le rôle des glycannes dans le repliement et l'assemblage des glycoprotéines E1 et E2. Nous avons montré que la glycosylation sur le site n°4 de la protéine E1 est un élément déterminant pour la formation d'un complexe E1E2 natif, alors que la mutation des sites 1, 2 ou 3 n'entraîne que peu ou pas de perturbations. Les hypothèses permettant d'expliquer l'importance de la glycosylation sur le site n°4 ont été testées. Enfin, nous avons testé la sécrétion de formes tronquées de la protéine E1, délétées leurs extrémités C-terminales. Nous avons pu mettre en évidence une forme tronquée très efficacement sécrétée. Une meilleure compréhension des mécanismes de formation des hétérodimères matures permettrait une optimisation de leur potentiel immunogénique. De même, des formes fortement sécrétées de la protéine E1 seraient très utiles pour constituer un outil diagnostique
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Spieker, Mark-Christoph [Verfasser]. "The origin of low-lying collective E1 and E2 strength in atomic nuclei / Mark-Christoph Spieker." München : Verlag Dr. Hut, 2017. http://d-nb.info/1126297844/34.
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Markson, Gabriel Benjamin. "A systematic analysis of the E1, E2, and E3 interactions within the human protein ubiquitination system." Thesis, University of Cambridge, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.612409.
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Ciczora, Yann. "Rôles fonctionnels des domaines transmembranaires des glycoprotéines d'enveloppe E1 et E2 du virus de l'hépatite C." Lille 2, 2006. http://www.theses.fr/2006LIL2S064.
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Le virus de l’hépatite C (VHC) code pour deux protéines d’enveloppe E1 et E2 associées sous forme d’hétérodimères. Les résidus chargés du domaine transmembranaire (TMD) de E2 (D728 et R730) n’ont pas la même contribution dans les différentes fonctions des TMDs. Bien que ces deux résidus soient nécessaires à la rétention de E2 dans le réticulum endoplasmique, l’acide aspartique est essentiel à la formation des hétérodimères. De plus, la mutation des résidus chargés du TMD de E2 altère l’infectiosité du virus. Nous avons effectué une mutagenèse de substitution tryptophane de chacun des résidus de ces domaines. Nous avons ainsi montré qu’en plus du résidu D728, les glycines 354 et 358 sont en partie responsables de la formation des hétérodimères E1E2. Enfin, nos observations indiquent que les TMDs de E1 et E2 sont impliqués dans l’entrée virale, et notamment dans une étape précoce de la fusion entre les membranes virale et cellulaireHepatitis C virus (HCV) encodes two envelope glycoproteins, E1 and E2 associated as heterodimers. These proteins are essential for virus infectivity. The two charged residues (Asp728 and Arg 730) of transmembrane domain (TMD) of E2 do not contribute equally in the glycoprotein functions. The two charged residues are required for ER retention, but only the aspartic acid is necessary for heterodimerization. Moreover the mutation of this charged residues affects the entry functions of these proteins. We have done a tryptophane scanning mutagenesis of each residue of these segments. The Asp728 and the two glycine residues (Gly354 and Gly358) are required for the formation of the heterodimer. Moreover other residues (Lys370, Leu726, Ala727, Ala729) are also implicated in these interactions. Finally, our observations indicate that the TMDs are also involved in virus entry. Indeed, some mutants of the TMDs of E1 and E2 affected an early step of the fusion between the viral and cell membrane
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Brett, Tricia Korrin. "The fate of estrone (E1), 17beta-estradiol (E2), estriol (E3) and 17alpha-ethinylestradiol (EE2) in surface waters." Thesis, University of British Columbia, 2014. http://hdl.handle.net/2429/46253.
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Lakes and rivers receiving wastewater treatment plant effluent contain many different endocrine disrupting compounds. Previous research into the fate of these compounds has focused on laboratory experiments that investigate a single scavenging mechanism, and there has been little research on the overall loss rate constants in receiving waters. This study evaluated the fate of estrone (E1), 17??-estradiol (E2), estriol (E3) and 17??-ethinylestradiol within three different receiving waters (a river, a large lake and a small reservoir) represented by two different mathematical models (plug flow reactor and continuously stirred tank reactor) and three different hydraulic residence times (<8 hours, >50 years and about 1 year). Wastewater treatment plant effluent samples and receiving waters were analysed for the four estrogens over a one year period. E1 and E2 were the only compounds detected and there was only enough data determine the fate of E1. A receiving water loss rate constant for E1 was calculated assuming first-order reaction kinetics. E1 loss was not detectable in the river and the large lake due to a very short and very long residence time, respectively. The time-weighted E1 loss rate constant within the small reservoir was found to be 0.0106 d-??. Data suggested that there may be a seasonal component to this loss rate that requires further investigation. The rate constant found suggests that E1 can be transported great distances within rivers.
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Soranzo, Thomas. "Approches Recombinantes pour l’Etude Structure/Fonction des Protéines E1, E2 et p7 du Virus de l’Hépatite C." Thesis, Université Grenoble Alpes (ComUE), 2015. http://www.theses.fr/2015GREAV056.
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Le virus de l'hépatite C (VHC) est une cause majeure d'affection hépatique chronique, notamment la cirrhose et le cancer du foie. On estime que 170 millions de personnes dans le monde sont des porteurs chroniques du VHC et que 3 à 4 millions de personnes sont infectées chaque année. Un des handicaps majeurs de la recherche sur le VHC est l'absence de systèmes de culture in vitro efficaces et de modèles animaux. Nous avons ainsi choisi une approche recombinante pour l'étude de protéines E1, E2 et p7 du VHC.Les protéines E1, E2 et p7 qui sont impliquées dans des étapes essentielles du cycle viral sont des protéines membranaires. Cependant, l'expression recombinante de cette classe de protéine est extrêmement complexe. En effet, la surexpression des protéines membranaires est souvent toxique pour les cellules hôtes. Ce phénomène est provoqué par l'agrégation ou la dégradation des protéines dans le cytoplasme dû à un manque de membrane disponible pour assurer leur intégration sur la cellule hôte. De plus, la surexpression de protéines membranaires induit la saturation de la machinerie cellulaire liée aux protéines membranaires. Ce détournement empêche le déroulement d'un cycle cellulaire normal et est ainsi fatal pour la cellule hôte. La forte concentration de protéines membranaires ou encore le fait que celles-ci soient hétérologues peut également provoquer la déstabilisation de la membrane de la cellule hôte et de son homéostasie. Afin de nous affranchir de ces limitations, nous avons utilisé une méthode de production des protéines membranaires sous forme native par un système acellulaire en présence de liposomes ; une technologie brevetée par l'université Joseph Fourier et exploitée par la société Synthelis. Dans un premier temps, nous avons procédé à la mise en place du système de production exploitant un lysat bactérien d'E. coli et d'un mélange énergétique complémentaire. Nous avons ensuite utilisé ce system pour étudier la viroporine p7. Cette protéine est essentielle pour la production de particules virales infectieuses et est impliquée dans l'assemblage viral ce qui en fait une cible thérapeutique intéressante. La production de protéoliposomes p7 en grande quantité nous a permis la caractérisation de la protéine par des techniques biochimiques et biophysiques. Nous avons mis en évidence l'inhibition de l'oligomérisation de p7 par le HMA qui ainsi inhibe sa fonction canal ionique. Grâce à la flexibilité du système d'expression acellulaire nous avons caractérisé la structure de la viroporine dans la membrane par réflectivité de neutron et avons confirmé la forme en entonnoir du complexe protéique. Des résultats préliminaires sur les proéoliposomes E1E2 quant à eux permettent d'espérer la production prochaine de particules virales mimant le VHC afin de mieux l'étudier et de lutter contre cette épidémie.L'ensemble de ces résultats confirment la pertinence de l'expression de protéines membranaires sous formes natives en système acellulaire en présence de liposomes. Les protéoliposomes produits constituent des nouveaux outils pour l'étude du VHC et permettent d'envisager de très grandes applications thérapeutiques ainsi que le développement de biomédicaments basés sur l'utilisation de protéines membranaires recombinantesThe Hepatitis C virus (HCV) is a major cause of chronic liver disease, including cirrhosis and liver cancer. An estimated 170 million people worldwide are chronically infected with HCV and 3 to 4 million people are infected each year. One of the major handicaps of the HCV research is the lack of effective in vitro culture systems and animal models. To adress this issue, we chose a recombinant approach to study the E1, E2 and p7 proteins of HCV.The E1, E2 and p7 proteins are involved in critical steps of the viral cycle. They are membrane proteins, a class of protein that is extremely complex to express. Indeed, overexpression of membrane proteins is often toxic to the host cells. This phenomenon is caused by protein aggregation or degradation in the cytoplasm due to a lack of available membrane space for their integration into the host cell. Moreover, overexpression of membrane proteins induces saturation of the cellular machinery linked to membrane proteins. This diversion prevents the flow of a normal cell cycle and is fatal to the host cell. Destabilization of the host cell's membrane and its homeostatis may also be caused by the high concentration of membrane proteins or their heterologous nature. To circumvent these limitations, we used a method for producing membrane proteins in their native form by a cell-free system in the presence of liposomes; a technology patented by the University Joseph Fourier and licenced by the startup company Synthelis. First, we have set up the cell-free production system using a bacterial lysate from E. coli and a complementary energy mix. We then used this system to study the p7 viroporine. This protein is essential for the production of infectious virus particles and is involved in viral assembly making it an attractive therapeutic target. The production of a large quantity of p7 proteoliposomes allowed us to characterize the protein by biochemical and biophysical techniques. We have demonstrated the inhibition of oligomerization of p7 by HMA, which thereby inhibits its ion channel function. Thanks to the flexibility of the cell-free expression system we have characterized the structure of the viroporine within the membrane in a neutron reflectivity assay and have confirmed the funnel shape of the protein complex. Preliminary results on proteoliposomes E1E2 offer hope for the production viral particles mimicking the hepatitis C virus in order to better study the virus and fight against this epidemic.Together, these results confirm the suitability of the expression of membrane proteins in native forms using a cell-free system in the presence of liposomes. Proteoliposomes products are a new tool for the study of HCV and consideration for very broad therapeutic applications and the development of biopharmaceuticals based on the use of recombinant membrane proteins
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Radtke, Christina [Verfasser], Birke Andrea [Akademischer Betreuer] Tews, Michael R. [Gutachter] Knittler, and Norbert [Gutachter] Tautz. "Charakterisierung der pestiviralen Glykoproteine E1 und E2 / Christina Radtke ; Gutachter: Michael Knittler, Norbert Tautz ; Betreuer: Birke Andrea Tews." Greifswald : Universität Greifswald, 2020. http://d-nb.info/1211087069/34.
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Radtke, Christina [Verfasser], Birke Andrea [Akademischer Betreuer] Tews, Michael [Gutachter] Knittler, and Norbert [Gutachter] Tautz. "Charakterisierung der pestiviralen Glykoproteine E1 und E2 / Christina Radtke ; Gutachter: Michael Knittler, Norbert Tautz ; Betreuer: Birke Andrea Tews." Greifswald : Universität Greifswald, 2020. http://d-nb.info/1211087069/34.
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Haddad, Juliano. "Etude biochimique et fonctionnelle de la glycoprotéine E1 du virus de l'Hépatite C (HCV)." Thesis, Lille 2, 2017. http://www.theses.fr/2017LIL2S029/document.
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Du fait de leur présence à la surface de la particule virale, les glycoprotéines d’enveloppe E1 et E2 du virus HCV jouent un rôle essentiel dans sa morphogenèse ainsi que lors de son entrée dans la cellule hôte. Jusqu’à récemment, les travaux de recherche sur les glycoprotéines d’enveloppe du virus HCV se sont essentiellement focalisés sur E2 car elle est la protéine d’attachement du virus. De plus, elle est la cible majeure des anticorps neutralisants et il a été longtemps postulé qu’elle était la protéine de fusion du virus. Cependant, les récentes publications de la structure de E2 ne mettent pas en évidence la présence d’un peptide de fusion et sa structure ne correspond pas aux critères attendus pour une protéine de fusion, suggérant que la glycoprotéine E1 seule ou en association avec E2 pourrait être responsable de l’étape de fusion. La structure de la région N-terminale de E1 (acides aminés 192 à 270) a récemment été résolue et a mis en évidence la présence d’une épingle à cheveux formée par 2 feuillets beta (β1 et β2) suivie par un segment de 16 acides aminés qui forme une hélice alpha (α1) flanquant 3 feuillets beta antiparallèles (β3, β4 et β5). En plus de la caractérisation de ces structures secondaires de E1, une région qui se situe au milieu de la protéine (approximativement entre les résidus 274 et 292) a été proposée avoir un rôle actif au cours du processus de fusion et elle pourrait correspondre à un peptide de fusion.Nous nous sommes basés sur ces travaux récents pour investiguer le rôle fonctionnel de la glycoprotéine E1 par une approche de mutagenèse dirigée des résidus conservés dans la région N-terminale et dans la région du potentiel peptide de fusion, dans le contexte d’un clone infectieux du HCV. Comme attendu, nos résultats indiquent que ces mutations introduites dans E1 n’ont aucun effet sur la réplication virale. Cependant, vingt-et-un parmi les vingt-huit mutants produits conduisent à une atténuation ou une perte de l’infectiosité virale. D’une manière très intéressante, deux mutants atténués, le T213A et le I262A, se sont montrés moins dépendants au co-récepteur claudine-1. D’autre part, nous avons montré que ces mutants utilisent un autre récepteur de la famille des claudines (claudine-6) pour l’entrée virale, indiquant ainsi un changement de dépendance à son co-récepteur claudine-1. A l’opposé, deux autres mutants, le L286A et le E303A, se sont révélés avoir une plus grande dépendance au co-récepteur claudin-1 pour l’entrée dans les cellules d’hépatome. Au cours de ce travail, nous avons également identifié une mutation intéressante à proximité du potentiel peptide de fusion. Cette mutation, G311A, conduit à la sécrétion de particules virales entières mais non infectieuses, suggérant un défaut d’entrée cellulaire pour ce virus. De façon très surprenante, nous avons également identifié une mutation (D263A) qui conduit à la sécrétion de particules virales dépourvues d’ARN génomique. Une caractérisation plus poussée de ce mutant a de plus révélé une modification dans la co-localisation subcellulaire entre l'ARN viral et la glycoprotéine E1, mettant en évidence pour la première fois un dialogue croisé entre E1 et l'ARN génomique du HCV lors de la morphogenèse du virus.En conclusion, nos observations permettent d’identifier précisément les régions spécifiques de la protéine E1 qui jouent un rôle dans l’assemblage et l’entrée du virus dans la cellule, mettant en évidence le rôle majeur de la glycoprotéine E1 au niveau des différentes étapes du cycle infectieux du HCVBeing part of the viral particle, HCV envelope glycoproteins E1 and E2 play an essential role in virion morphogenesis as well as in HCV entry into liver cells. These glycoproteins form a non-covalent heterodimer, and until recently, research on HCV envelope glycoproteins has been mainly focused on E2. Indeed, this glycoprotein is the receptor-binding protein, it is also the major target of neutralizing antibodies and it was postulated to be the fusion protein. However, the recent publications of the structure of E2 do not show the presence of a fusion peptide and its structure does not fit with what one would expect for a fusion protein, suggesting that E1 alone or in association with E2 might be responsible for the fusion step. Concerning E1, only the crystal structure of the two-fifth N-terminal region, comprising amino acids 192 to 270, has been reported. This partial structure reveals a complex network of covalently linked, intertwined homodimers. The overall fold of the N-terminal E1 monomer consists of a beta-hairpin (β1 and β2) followed by a segment composed of a 16 amino-acid long alpha-helix (α1) flanking a three-strand antiparallel beta-sheet (β3, β4 and β5). In addition to the characterization of secondary structures within E1, a region located in the middle of the polypeptide (approximately between aa 274 and 292) has been suggested to play an active role during the fusion process and might potentially act as a fusion peptide. We took advantage of these recently published data to further investigate the functional role of HCV glycoprotein E1 by using a site-directed mutagenesis approach targeting conserved amino acids in the N-terminal region as well as in the region postulated to contain the fusion peptide in the context of an infectious clone. As expected, our results indicate that these mutations have no effect on virus replication. However, twenty-one out of twenty-eight mutations led to attenuation or inactivation of infectivity. Interestingly, two attenuated mutants, T213A and I262A, were less dependent on tight junction protein claudin-1, a co-receptor for HCV. Instead, these mutant viruses relied on another claudin (claudin-6) for cellular entry, indicating a shift in receptor dependence. In contrast, two other mutants, L286 and E303, were more dependent on claudin-1 for cellular entry into hepatoma cells cells. We also identified an interesting mutation downstream of the putative fusion peptide, G311A, which leads to the release of non-infectious particles having a defect in cellular entry. Finally, an unexpected phenotype was also observed for D263A mutant, which was no longer infectious but led to the secretion of viral particles devoid of genomic RNA. Further characterization of the D263A mutant revealed a change in subcellular co-localization between HCV RNA and E1, highlighting for the first time a crosstalk between HCV glycoprotein E1 and the genomic RNA during HCV morphogenesis.In conclusion, our observations allowed for the identification of specific regions in the E1 glycoprotein that play a role in virion assembly and entry, highlighting the major role played by this protein at different steps of the HCV infectious cycle
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Liu, Bing. "Structure insights into the autoinhibitory mechanism of the deubiquitinating enzyme USP25 and into the SUMO E1-E2 protein-protein recognition." Doctoral thesis, Universitat Autònoma de Barcelona, 2018. http://hdl.handle.net/10803/665314.
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La ubiquitinació i la SUMOilació són dos de les modificacions post-traduccionals més estudiades (PTM). En aquesta tesi, ens hem centrat en estudiar USP25, USP28 i en el reconeixement proteïna-proteïna dels enzims SUMO E1-E2 d’aquestes dues vies PTM. USP25 i USP28 són de-ubiquitinases que tenen papers importants en processos cel·lulars i les seves activitats enzimàtiques estan regulades per diversos PTMs, incloent SUMOilació, ubiquitinació i fosforilació. La interacció proteïna-proteïna de SUMO E1-E2 és un pas essencial de discriminació en la via de la conjugació. En aquesta tesi, els principals objectius inclouen l'elucidació de les bases estructurals de la regulació de l'activitat de USP25 i USP28, així com poder desxifrar els determinants estructurals en l'especificitat proporcionada per la interacció SUMO E1-E2.
Hem resolt l'estructura cristal·lina de la USP25 humana (18 - 714). Sorprenentment, USP25 mostra una estructura quaternària homotetramèrica que està directament implicat en la inhibició de la seva activitat enzimàtica, i revela un nou mecanisme de tetramerització/inhibició. Assaigs bioquímics i cinètics in vitro amb construccions de dímer, tetràmer de USP25 donen suport a aquest mecanisme, mostrant en tots els casos una major activitat catalítica en el dímer. A més, la forta estabilització de les tankirases en cèl·lules en cultiu mitjançant l'expressió ectòpica del dímer de USP25 verifica la rellevància biològica d'aquest nou mecanisme de tetramerització/inhibició.
Pel que fa a la interacció E1 UFD - E2, hem resolt l'estructura cristal·lina del complex E1 UFD - E2 tant en humans com en A. thaliana. Tot i la baixa homologia de seqüència presentada en la superfície d’interacció de UFD entre especies, la comparació estructural entre complexos revela determinants comuns en les interfícies entre humans, llevats i A. thaliana. La comparació estructural també revela una forta conservació en la superfície d’interacció de les E2 entre espècies, tot i la forta especificitat mostrada en assajos de conjugació de SUMO en a cada organisme. Curiosament, hem trobat residus de la E2 fora de la superfície d’interacció amb UFD que tenen un impacte en la conjugació de SUMO, el que suggereix la presència d’altres determinants estructurals diferents als de la superfície d’interacció entre E2 i E1 UFD en l'especificitat de la conjugació de SUMO.Ubiquitination and SUMOylation are of the most studied post-translational modifications (PTMs). Here, we focus on USP25, USP28, and the SUMO E1-E2 protein-protein recognition in these two PTM pathways. USP25 and USP28 have important roles in cellular processes, and their enzymatic activities are regulated by diverse PTMs including SUMOylation, ubiquitination, and phosphorylation. SUMO E1-E2 protein-protein interaction is a major discrimination step in the conjugation pathway. In this thesis, the main goals include the elucidation of the structural basis for the activity regulation of USP25 and USP28, as well as to decipher the structural determinants for the specificity provided by the E1 UFD-E2 interaction. We have solved the crystal structure of human USP25 (18 – 714). Unexpectedly, USP25 displays a homotetrameric quaternary assembly that is directly involved in the inhibition of its enzymatic activity, revealing a novel tetramerization/inhibition mechanism. In vitro biochemical and kinetic assays with dimer, tetramer and truncation constructs of USP25 support this mechanism, displaying in all cases a higher catalytic activity in the dimer assembly. Moreover, the strong stabilization of tankyrases in cultured cells by the ectopic expression of the USP25 dimer verifies the biological relevance of this novel tetramerization/inhibition mechanism. Regarding to the E1 UFD-E2 interaction, we have solved the crystal structure of the E1 UFD-E2 complex in both human and A. thaliana. Despite the low sequence homology displayed by the UFD binding interface, structural comparison between complexes reveals common determinants in the interfaces between human, yeast, and A. thaliana. Structural comparison also reveals a strong conservation in the E2 binding interface across species, despite the strong specificity displayed in SUMO conjugation assays for each organism. Interestingly, E2 residues outside the UFD interface had impact on SUMO conjugation, suggesting the contribution of determinants other than the primary UFD binding interface in the specificity of the conjugation system.
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Stockhammer, Engelbert, and Özlem Onaran. "Accumulation, distribution and employment. A structural VAR approach to a Post-Keynesian Macro Model." Inst. für Volkswirtschaftstheorie und -politik, WU Vienna University of Economics and Business, 2002. http://epub.wu.ac.at/1220/1/document.pdf.
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The paper investigates the relation between effective demand, income distribution and unemployment empirically. Its aim is to evaluate Keynesian, Kaldorian and neoclassical hypotheses about the determination of labor market variables. To do so, a vector autoregression model consisting of capital accumulation, capacity utilization, the profit share, unemployment and the growth of labor productivity is estimated. A general post-Keynesian model following the lines of Kalecki and Kaldor is presented and provides the specification for a structural VAR. The model is estimated for the USA, UK and France. (authors' abstract)Series: Working Papers Series "Growth and Employment in Europe: Sustainability and Competitiveness"
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Laurans, Françoise. "Definition histologique et approche moleculaire des interactions peupliers/rouilles (populus deltoides x p. Nigra cv. Ogy/melampsora larici-populina races e1 et e2)." Orléans, 1997. http://www.theses.fr/1997ORLE2017.
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Ce travail a pour objectif la caracterisation physiologique et moleculaire des interactions populus/melampsora et vise a la comprehension du determinisme genetique des mecanismes de defense des peupliers a l'attaque des rouilles. La definition spatio-temporelle des mecanismes de defense qui interviennent de maniere preponderante dans les deux type d'interaction (compatible et incompatible) d'un meme systeme hote/pathogene (populus deltoides x p. Nigra cv. Ogy/melampsora larici-populina race e1 ou e2) a ete abordee. Deux approches ont ete conduites parallelement. D'une part, une approche histologique a permis d'etablir la cinetique d'infection et de developpement du champignon pathogene melampsora larici-populina au sein des tissus foliaires de peuplier cultivar ogy. L'etude comparee de deux interactions, compatible et incompatible, a mis en evidence un certain nombre d'evenements histologiques precis permettant de differencier ces deux interactions. D'autre part, une approche moleculaire par analyses en northern blot et hybridation in situ a permis d'acceder a l'etude spatio temporelle de l'expression de differents genes de defense.
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Onaran, Özlem, and Engelbert Stockhammer. "Do profits affect investment and employment? An empirical test based on the Bhaduri-Marglin model." Inst. für Volkswirtschaftstheorie und -politik, WU Vienna University of Economics and Business, 2005. http://epub.wu.ac.at/1534/1/document.pdf.
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In this study, a Kaleckian-Post-Keynesian macroeconomic model, which is an extended version of the Bhaduri and Marglin (1990) model, serves as the starting point. The merit of a Kaleckian model for our purposes is that it highlights the dual function of wages as a component of aggregate demand as well as a cost item as opposed to the mainstream economics, which perceive wages merely as a cost item. Depending on the relative magnitude of these two effects, Kaleckian models distinguish between profit-led and wage-led regimes, where the latter is defined as a low rate of accumulation being caused by a high profit share. Are actual economies wage-led or profit-led? Current orthodoxy implicitly assumes that they are profit-led, and thus supports the neoliberal policy agenda. The purpose of the paper is to carry this discussion into the empirical terrain, and to test whether accumulation and employment are profit-led in two groups of countries. We do so by means of a structural vector autoregression (VAR) model. The model is estimated for USA, UK and France to represent the major developed countries, and for Turkey and Korea to represent developing countries. The latter are chosen since they represent two different export-oriented growth experiences. The results of the adjustment experiences of both countries are in striking contrast to orthodox theory, however they also present counter-examples to each other in terms of their ways of integrating into the world economy. (author's abstract)Series: Working Papers Series "Growth and Employment in Europe: Sustainability and Competitiveness"
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Uribe, Laplechade Catalina del Carmen. "Evaluación de la aplicación de estiércol animal en relación a la presencia, disponibilidad y biodisponibilidad de estrona (E1), 17β-estradiol (E2) y 17α-etinilestradiol (EE2) en suelos degradados." Tesis, Universidad de Chile, 2018. http://repositorio.uchile.cl/handle/2250/151314.
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Tesis presentada a la Universidad de Chile para optar al grado de Magíster en Química área de Especialización en Química Ambiental y Memoria para optar al Título de QuímicoCon el objeto de buscar alternativas de producción más amigables con el medio ambiente, y evitar el deterioro de los ecosistemas, la producción silvoagropecuaria ha generado opciones sustentables y ecológicas como la agricultura orgánica, la cual conserva o aumenta la materia orgánica del suelo reciclando los residuos de cosecha, poda, estiércol y guano de animales, a través de distintos sistemas de incorporación al suelo. Según la Ley 20.089, el estiércol corresponde a fecas, orinas y productos de cama de animales, que no han sido compostado. El guano y la orina de los animales contienen cantidades importantes de nitrógeno, fósforo, potasio y otros elementos necesarios para el crecimiento de las plantas. La combinación de estiércol, paja de cereales y restos hortícolas es una mezcla de alta calidad como abono para el suelo. La composición de los guanos es muy variable y generalmente depende de la dieta que se le suministra al animal. La legislación chilena señala que la carga ganadera establecida se debe fijar considerando que no se debe sobrepasar el límite de 170 kg de nitrógeno·ha-1·año-1. Por otro lado, la ganadería intensiva crea problemas de depósito de estiércol y contaminación de agua. En este tipo de ganadería de producción intensiva el uso de fármacos u otros insumos es una práctica habitual que tiene como objetivos, aumentar la eficiencia en la engorda de los animales y la producción de leche. Uno de los problemas que puede provocar este hecho, es el aumento de la concentración de algunos contaminantes emergentes como lo son las hormonas estrogénicas naturales y sintéticas, por ejemplo, la estrona (E1), el 17β-estradiol (E2) y el 17α-etinilestradiol (EE2), que están consideradas disruptores endocrinos, y aunque se encuentren en muy bajas concentraciones, pueden provocar serios daños en animales y en seres humanos. Por esto, el objetivo de este trabajo fue evaluar la biodisponibilidad de dos hormonas naturales (E1 y E2) y una hormona sintética (EE2) en suelos restaurados con estiércol, empleando plantas de trigo como indicadores y compararlas con una extracción mediante hidroxipropil-β-ciclodextrina (HPCD). Para esto se evaluó la concentración de los tres analitos (E1, E2 y EE2) en estiércol de vacuno, cerdo y caballo, realizando una extracción asistida con ultrasonido, seguido de una etapa de clean up, derivatización y cuantificación en un cromatógrafo de gases acoplado a espectrometría de masas. También se realizó una caracterización física y química de los tres tipos de suelos y los distintos estiércoles, se determinó la fracción biodisponible en plantas de trigo cultivadas en suelos enmendados con estiércol y se estimó la concentración biodisponible mediante la extracción de las hormonas con ciclodextrina, para así validar este método como biosimulador a través de la comparación con los resultados obtenidos en el bioensayo con plantas de trigo. Finalmente, se realizó un estudio de degradación de las tres hormonas estrogénicas, aplicadas directamente a los suelos o agregadas a través del estiércol enriquecido con éstas, en dos periodos de tiempo, el primero durante treinta días y el segundo en siete días. Las hormonas naturales E1 y E2 se encontraron en los tres estiércoles, mientras que solo en el estiércol de cerdo se encontró la hormona sintética EE2. Los tres analitos se encontraron biodisponibles en las raíces de las plantas de trigo, siendo la EE2 en el suelo Codigua la que presentó mayor biodisponibilidad, el bioensayo con ciclodextrina correlacionó con la fracción biodisponible en las plantas de trigo, por lo tanto, el método biosimulador serviría como sistema predictivo en los tres suelos utilizados. Finalmente, el tiempo de degradación de las hormonas fue en general rápido, las tres hormonas se degradaron en su mayoría dentro de los dos primeros días, siendo la EE2 la más estable y la E2 la menos estable, ya que posiblemente ocurriría su degradación por oxidación a E1, por lo tanto, la concentración de E1 en las primeras horas tendería a aumentar
Intending to find environmentally production alternatives and avoid ecosystem damage, is that agro forestry production has generated sustainable and ecological alternatives as organic agriculture, which preserves or increases organic material from the ground by recycling harvest, pruning, manure and animal guano remains, all these through different ground incorporation systems. According to law no. 20.089, manure is composed by non composted feces, urine and animal bed products. Guano and urine from animals contains a significant amount of nitrogen, phosphor, potassium and some other elements needed for plants growth. The mixture made of manure, cereal straw and horticultural remains is a high quality fertilizer for the soil. Different guano compositions is very variable y generally depends on the animal diet. Current legislation points that established livestock load must be determinated considering not exceeding 170 kilograms limit of nitrogen·ha-1·year-1. On the other hand, intensive animal breeding creates manure deposit and water pollution problems. In this type of intensive animal breeding production, it is usual practice the use of drugs and other supplies that look for increase efficiency on animal fattening and milk production. One of the problems that this fact could cause is the increase on some emerging pollutants concentration as natural and synthetic estrogen hormones, like estrone (E1), 17β-estradiol (E2) y 17β-ethinylestradiol (EE2), which are considered as endocrine disruptors, that although are found on low concentration, can cause serious damage both in animals and in humans. Therefore, the main objective of this work was to evaluate the effect of the application of manure on soils and determine the bioavailable fraction of two natural hormones (E1 and E2) and a synthetic hormone (EE2) in wheat plants grown in the soils restored with manure, comparing the results with a bioassay using hydroxypropyl-β-cyclodextrin (HPCD) as an extractant. For this, the concentration of the three analytes (E1, E2 and EE2) in cow, pig and horse manure was evaluated, performing an assisted extraction with ultrasound, then a clean up, derivatization and measuring were made by in a coupled gas chromatograph to mass spectroscopy. A physical and chemical characterization of the three types of soils and the different manure was also carried out, the bioavailable fraction in wheat plants cultivated in soils amended with manure was determined and the bioavailable concentration was estimated by extracting the hormones with cyclodextrin, to validate this method as a biosimulator through the comparison with the results obtained in the bioassay with wheat plants. Finally, a study of degradation of the three estrogenic hormones, applied directly in the soil or added through the manure enriched with these, in two periods of time, the first during a month and the second for seven days. The natural hormones E1 and E2 were found in the three manures, and only synthetic mannitol EE2 was found in pig manure. The three analytes were found bioavailable in the roots of wheat plants with EE2 in Codigua soil having the highest bioavailability, the bioassay with cyclodextrin correlated with the bioavailable fraction in wheat plants, validating the biosimulator method as a predictive method, and finally, the time of degradation of hormones is generally fast, the three hormones are degraded mostly within the first two days, EE2 being the most stable and E2 the least stable, may because of this is degraded by oxidation to E1, therefore the concentration of E1 in the first hours tends to increase
Fondecyt
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Greenlees, Paul Thomas. "Identification of excited states and evidence for octupole feformation in '2'2'6U." Thesis, University of Liverpool, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.367435.
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Amorim, Kamila Pereira de. "Desenvolvimento de um método por ponto nuvem dos hormônios naturais E1 e E2 em amostras de urina e determinação por CLAE/EC utilizando eletrodo de diamante dopado com boro." Universidade Federal de Goiás, 2015. http://repositorio.bc.ufg.br/tede/handle/tede/5073.
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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES
Cloud point extraction method (CPE) was used for the determination of estrone (E1) and 17β-estradiol (E2) hormones in human urine. The combination of the electrochemical detection techniques with high-performance liquid chromatography (HPLC-EC) was used for the detection and quantification of these hormones. A boron doped diamond electrode (BDD) pretreated cathodically was used as electrode material for all electrochemical measurements. The optimized chromatographic parameters resulted in a mobile phase composition of KH2PO4 (0.01 mol L-1; pH 5.0) / ACN (72:28 V/V), flow 1.2 ml min-1. An applied potential for electrochemical detection of 1.0 V x Ag/AgCl (3.0 mol L-1) was selected from hydrodynamic voltammograms constructed for each hormone changing the potential between 0.3 V and 1.2 V x Ag/AgCl (3.0 mol L-1). Limits of detection (S/N = 3) of 500 ng mL-1 and limits of quantification of 800 ng mL-1 were obtained for both E1 and E2 hormones without any extraction process. Urine samples at pH 5.0 and 7.0 were investigated aiming the influence of pH on the efficiency of the CPE process, and the optimum results for the most current signal of the hormones was obtained at pH 7.0. Extractor solvent volumes were changed in the 0.5-2.5 mL range, and the optimum results were obtained when using 1.0 mL of Tergitol TMN-6 surfactant (10% aqueous solution). From the equation of the calibration curves obtained with and without the CPE procedure it was possible to determine the pre-concentration factor (FC) and all the other parameters involving the efficiency of CPE method. A comparison of the efficiency of CPE method with direct liquid-liquid extraction with the organic solvent CCl4 was carried out and the results showed that the CPE method was quite superior to liquid-liquid extraction. The validation of the method was carried out from intra-day recovery experiments and inter-day and evaluated the accuracy, precision and repeatability. The proposed method was applied to individual samples of urine of 1 man, 1 pregnant woman, 1 woman in fertile age, and 1woman in lactating stage. The values of the variation coefficients of the recovery percentages were lower than 15%.
Método de extração por ponto nuvem (EPN) foi usado para a determinação dos hormônios estrona (E1) e 17β-estradiol (E2) em urina humana. A combinação entre as técnicas de detecção eletroquímica e cromatografia líquida de alta eficiência (CLAE-EC) foi usada para a detecção e quantificação desses hormônios. Um eletrodo de diamante dopado com boro (DDB) pré-tratado catodicamente foi usado como material de eletrodo para todas as determinações eletroquímicas. Os parâmetros cromatográficos otimizados resultaram em uma composição de fase móvel de KH2PO4 (0,01 mol L-1; pH 5,0) / ACN (72:28 V/V) vazão de 1,2 mL min-1. Um potencial aplicado para a detecção eletroquímica de 1,0 V x Ag/AgCl (3 mol L-1) foi selecionado a partir de voltamogramas hidrodinâmicos construídos para cada hormônio variando-se o potencial entre 0,3 V e 1,2 V x Ag/AgCl (3,0 mol L-1). Limites de detecção (S/R = 3) de 500 ng mL-1 e limites de quantificação de 800 ng mL-1 foram obtidos para ambos os padrões dos hormônios E2 e E1 sem qualquer processo de extração. Amostras de urina em pH 5,0 e 7,0 foram investigadas quanto a influência do pH na eficiência do processo de extração, e o melhor resultado referente ao maior sinal de corrente dos hormônios foi obtido em pH 7,0. Os volumes de solvente extrator foram variados na faixa de 0,5-2,5 mL e o melhor resultado referente ao sinal de corrente dos hormônios foi obtido pelo uso de 1,0 mL de solução aquosa 10% do surfactante Tergitol TMN-6. A partir da equação da reta obtida das curvas analíticas, com e sem o procedimento de EPN, foi possível determinar o fator de pré-concentração (FC) e todos os demais parâmetros envolvendo a eficiência do método de EPN. Uma comparação sobre a eficiência dos métodos de EPN com extração direta líquido-líquido com o solvente orgânico CCl4 foi realizada e os resultados mostraram que método de EPN mostrou-se bastante superior a extração líquido-líquido. A validação do método foi feita a partir de ensaios de recuperação intra-dia e inter-dia, sendo avaliadas a exatidão, precisão e repetitividade. O método proposto foi aplicado em amostras individuais de urina de 1 homem, 1 mulher gestante, 1 mulher em idade fértil e 1 mulher lactante. Os valores dos coeficientes de variação das porcentagens de recuperação foram menores que 15%.
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Moenne-Loccoz, Rémy. "Impact des glycoprotéines d'enveloppe E1 et E2 du virus de l'hépatite C sur la réponse au traitement antiviral interféron-a pégylé/ribavirine chez des patients atteints d'hépatite chronique C." Strasbourg, 2011. https://publication-theses.unistra.fr/public/theses_doctorat/2011/MOENNE-LOCCOZ_Remy_2011.pdf.
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Le traitement de référence composé d’interféron-alpha pégylé/ribavirine n’est efficace que chez 50% des patients chroniquement infectés par un virus de l’hépatite C (VHC)-génotype 1. La haute variabilité des glycoprotéines d’enveloppe E1/E2 du VHC pourrait indirectement contribuer à la résistance virale au traitement par la sélection de souches avec un potentiel infectieux (p. I) élevé et/ou une capacité accrue à échapper à l’immunité. Cette hypothèse a été évaluée in silico puis par analyses fonctionnelles in vitro (pseudoparticules HCVpp). Des signatures moléculaires (SM) et des réseaux minimaux d’acides aminés (aa) covariants, définis sur E1/E2, étaient corrélés à la réponse au traitement. Trois des quatre SM définies et fonctionnellement évaluées montraient des résultats en accord avec l’hypothèse. Les résidus 431A et 642V liés à la non-réponse (NR) entraînaient une diminution de la neutralisation des HCVpp par les anticorps (Ac) circulant dans le sérum de patients, une augmentation 431A- ou 642V-dépendante du p. I. Des HCVpp à l’étape d’entrée dans les cellules, et une interaction augmentée avec CD81 et SR-BI via 431A. Le résidu 219T lié à la réponse (R) diminuait le p. I des HCVpp. Des réseaux d’aa covariants séparaient les souches NR des souches R et comportaient 3 des 4 SM citées plus haut. Conclusion : Nos résultats sont en faveur d’une contribution indirecte de E1/E2 du VHC à l’efficacité du traitement. Les résidus 431A et 642V (NR) favorisent le p. I des HCVpp et un échappement aux Ac neutralisants alors que le résidu 219T (R) diminue le p. I, suggérant que les interactions virus-hôte durant l’entrée virale peuvent intervenir dans l’échec du traitement anti-VHCThe standard of care (SOC) treatment, i. E. Pegylated interferon-alpha/ribavirin, is efficient in only 50% of patients chronically infected with hepatitis C virus (HCV)-genotype 1. The high variability of HCV E1/E2 envelope glycoproteins may indirectly contribute to viral resistance to treatment by selection of strains with high infectivity and/or increased ability to escape to immunity. This hypothesis was investigated by in silico and in vitro functional analyses (pseudoparticles HCVpp). Molecular signatures (MS) and covariant amino acid minimal networks, defined on E1/E2, were correlated with treatment response. Three out of the four MS which were defined and functionally assessed showed results concordant with the hypothesis. The nonresponse (NR)-related residues 431A and 642V led to a decrease in antibody (Ab)-mediated HCVpp neutralization using patients sera, a 431A or 642V-dependent increase of HCVpp infectivity at the entry step, and a 431A-dependent increase of interaction with CD81 and SR-BI. The response (R)-related residue 219T decreased HCVpp infectivity. Minimal networks of covariant amino acids separated NR-related from R-related strains and included three out of the four MS previously mentioned. Conclusion: Our results support an indirect contribution of HCV E1/E2 to treatment efficacy. NR-related 431A and 642V favour HCVpp infectivity with concomitant escape from neutralizing Ab while R-related 219T decreases HCVpp infectivity, suggesting that virus-host interactions during viral entry may be involved in the SOC treatment failure
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Guimarães, Tatiane Sant\'Ana. "Detecção e quantificação dos hormônios sexuais 17 \'beta\'-estradiol (E2), estriol (E3), estrona (E1) e 17 \'alfa\'-etinilestradiol (EE2) em água de abastecimento: estudo de caso da cidade de São Carlos, com vistas ao saneamento ambiental." Universidade de São Paulo, 2008. http://www.teses.usp.br/teses/disponiveis/18/18138/tde-26022009-100015/.
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Um dos grandes problemas da engenharia ambiental é a contaminação dos corpos hídricos. Os sanitaristas têm se preocupado com os hormônios sexuais, notadamente os estrógenos, compostos extremamente ativos biologicamente, que têm sido referidos como agentes etiológicos de feminilização e de vários tipos de cânceres. Os estrógenos naturais 17 \'beta\'-estradiol (E2), estriol (E3), estrona (E1) e o sintético 17 \'alfa\'-etinilestradiol (EE2), desenvolvidos para uso médico em terapias de reposição hormonal feminina e métodos contraceptivos, são os que despertam maiores preocupações, pela contínua introdução ao ambiente; hormônios que possuem a melhor conformação reconhecida pelos receptores que resultam respostas máximas, por isso são considerados responsáveis pela maioria dos efeitos disruptores desencadeados pela disposição de efluentes. A mudança de padrões quanto à atividade sexual dos jovens e a preocupação com o planejamento familiar, levaram ao grande consumo de contraceptivos que, através da urina, são levados pela rede coletora aos corpos de água. O indiscriminado uso desses hormônios na bovinocultura, suinocultura, avicultura e aqüicultura são responsáveis por parte considerável desse contaminante nos mananciais. Os hormônios excretados através da urina e fezes e agentes provenientes das indústrias de processamento de alimentos preocupam os sanitaristas porque o lançamento de efluentes in natura ou tratados, são as principais vias de contaminação do ambiente aquático, quer pelo déficit de infra-estrutura em saneamento, quer pela ineficiência tecnológica e/ou operacional na remoção desses compostos nas estações de tratamento de água ou de efluentes. Apesar de possuírem meia-vida relativamente curta, quando comparados a outros compostos orgânicos como praguicidas, os estrógenos naturais são continuamente introduzidos no ambiente, o que lhes confere caráter cumulativo. A proposta desta pesquisa foi verificar a presença de estrógenos na água bruta que chega à Estação de Tratamento de Água, após seu tratamento, em água tratada por osmose reversa e por tecnologia Milli Q (marca registrada). Para verificar e quantificar presença desses hormônios estrógenos em água de abastecimento de São Carlos-SP, foram realizadas exames através de imunoensaio quimiluminescente e radioimunoensaios. Os resultados apontaram que a ETA não possui solução eficiente para a remoção dos analitos de interesse dessa pesquisa, uma vez que na água tratada foram encontrados valores semelhantes aos da água bruta.One of the major problems of environmental engineering is the water contamination. The sanitary persons have been concerned with the gonadal hormones, notably the estrogen, biologically active compounds extremely, which have been referred to as etiologic agents of feminization and several types of cancers. The natural estrogen 17 \'beta\'-estradiol (E2), estriol (E3), estrone (E1) and synthetic 17 \'alpha\'-ethinylestradiol (EE2), developed for medical use of hormone replacement therapy in women and contraceptive methods, are those that attract larger concerns by the continuous introduction into the environment; hormones that have the best conformation recognized by receptors that result answers maximum, so they are considered responsible for most of the effects disruptors triggered by the wastewater disposal. The change in patterns on the sexual activity of young people and the concern with family planning, led to the large consumption of contraceptives that, in the urine, are led by the distribution net to water. The indiscriminate use of these hormones in cattle, pigs, poultry and aquaculture are responsible for part of this contaminant in the source. The hormones excreted in the urine and feces and agents from the food processing industries in the sanitary concern that the launch of effluents in nature or treated, are the main routes of contamination of the aquatic environment, either by lack of infrastructure, sanitation, or by inefficiency technological and/or operating in the removal of these compounds in the treatment plants, water or effluent. Despite having relatively short stocking-lige when compared to other organics such as pesticides, natural estrogens are continuously released into the environment, which gives them character cumulative. The proposal of this research was to verify the presence of estrogen in the gross water that arrives at the water treatment plant, after the treatment, in water treated by reverse osmosis and by Milli Q technology. To verify and quantify presence of these hormones estrogen in water supply of San Carlos-SP, examinations were conducted through immunoassay chemiluminescent and radioimunoassays. The results showed that the ETA has no efficient solution for removal of interest analytes of this research, because in the treated water were found values similar to crude water.
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Morice, Yoann. "Le virus de l'hépatite C : études de la variabilité inter- et intra-génomique : production sous forme soluble de protéines membranaires impliquées dans l'interaction virus-cellule hôte." Paris 7, 2002. http://www.theses.fr/2002PA077217.
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Klein, Marina [Verfasser], Michael [Akademischer Betreuer] Roggendorf, Elke [Akademischer Betreuer] Cario, and Daniel [Akademischer Betreuer] Hoffmann. "Evolution of the envelope proteins E1 and E2 and of specific humoral immune response to these proteins in a group of patients infected by HCV in a single-source outbreak / Marina Klein. Gutachter: Elke Cario ; Daniel Hoffmann. Betreuer: Michael Roggendorf." Duisburg, 2011. http://d-nb.info/1015361803/34.
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Steiner, Laure D. "A Study of the fate and transport of estrogenic hormones in dairy effluent applied to pasture soils." Diss., Lincoln University, 2009. http://hdl.handle.net/10182/1306.
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The disposal of waste from agricultural activities has been recognised as a source of environmental contamination by endocrine disrupting chemicals (EDCs). The New Zealand dairy industry produces a large volume of dairy farm effluent, which contains EDCs in the form of estrogens. Most of this dairy farm effluent is applied onto the land for disposal. Groundwater and soil contamination by estrogens following waste application on the land have been reported overseas, but our understanding of the processes and factors governing the fate of estrogens in the soil is poor. Therefore the main goal of the present study was to better understand the fate and transport of estrogens, in particular 17β-estradiol (E2) and estrone (E1) in soil. In order to quantify E1 and E2 in drainage water and soil samples, chemical analysis by gas-chromatography mass-spectrometry (GC-MS) was carried out. This included sample extraction, sample clean-up through silica gel and gel permeation chromatography, and sample extract derivatisation prior to analysis. In order to develop a reliable method to extract estrogens from soil, research was conducted to optimise E1 and E2 extraction conditions by adjusting the number of sonication and shaking events, as well as the volume and type of solvent. Among five solvents and solvent mixtures tested, the best recovery on spiked and aged soil was obtained using an isopropanol/water (1:1) mix. A microcosm experiment was carried out to determine the dissipation rates of E2 and E1, at 8°C and at field capacity, in the Templeton soil sampled at two different depths (5-10 cm and 30-35 cm). The dissipation rates decreased with time and half-life values of 0.6-0.8 d for E1 and 0.3-0.4 d for E2 were found for the two depths studied. A field transport experiment was also carried out in winter, over three months, by applying dairy farm effluent spiked with estrogens onto undisturbed Templeton soil lysimeters (50 cm in diameter and 70 cm deep). The hormones were applied in dairy farm effluent at 120 mg m⁻² for E2 and 137 mg m⁻² for E1. The results of the transport experiment showed that in the presence of preferential/macropore flow pathways 0.3-0.7% of E2 and 8-13% of E1 was recovered in the leachate at the bottom of the lysimeters after 3 months, and 1-7% of the recovered E2 and 3-54% of the recovered E1 was leached within 2 days of application. These results suggest that leaching of estrogens via preferential/macropore flow pathways is the greatest concern for groundwater contamination. In the absence of preferential/macropore flow pathways, a significant amount (> 99.94%) of both hormones dissipated in the top 70 cm of soil, due to sorption and rapid biodegradation. Surprisingly, in all cases, estrogen breakthrough occurred before that of an inert tracer (bromide). This could not be explained by the advection-dispersion transport of estrogens, nor by their presence as antecedent concentrations in the soil. It was therefore suggested that colloidal enhanced transport of estrogens was responsible for the earlier breakthrough of estrogens and caused the leaching of a fraction of the applied estrogens to a soil depth of 70 cm. A two-phase model, adapted from a state-space mixing cell model, was built to describe the observed estrogen transport processes under transient flow. The model takes into account 3 transport processes namely, advection-dispersion, preferential/macropore flow and colloidal enhanced transport. This model was able to successfully describe the estrogen transport observed from the lysimeters.
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Tsai, Chia-hao, and 蔡佳豪. "Structural Characterization of HCV E1 and E2 Proteins." Thesis, 2008. http://ndltd.ncl.edu.tw/handle/32252340507340326948.
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碩士慈濟大學
分子生物及細胞生物研究所
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Abstract Hepatitis C virus (HCV) is an enveloped, positive-stranded RNA virus classified in the Hepacivirus genus of the Flaviviridae family. The HCV genome encodes three structural proteins: a capsid protein and two envelope glycoproteins, E1 and E2. E1 and E2 are thought to play pivotal roles at different steps of the HCV replicative cycle. There is now strong evidence that E1 and E2 are essential for host-cell entry, binding to receptor(s), inducing fusion with the host-cell membrane as well as assembling viral particle. E1 and E2 are type I transmembrane (TM) glycoproteins, with an N-terminal ectodomain and short C-terminal TM domain. These proteins interact with each other and assemble as noncovalent heterodimers. Like other viral envelope proteins involved in host-cell entry, HCV envelope proteins are thought to induce fusion between the viral envelope and a host-cell membrane. The HCV envelope glycoproteins E1 and E2 are thought to be class II fusion proteins because the putative fusion peptide is proposed to localize in an internal sequence linked by antiparallel β-sheets. We have prepared several constructs containing truncated E1 or E2. The truncated E1 or E2 protein can be expressed and purified from E. coli. Circular dichroism analysis of these expressed proteins showed little difference between pH 7 and 6. Only E1192~260 fragment (amino acid sequence 192 to 260) could induce liposome fusion at low pH, while other fragments could not. Thus, our data suggest that amino acid between sequence 232~260 play an role in the event of membrane fusion.
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Whitehurst, Christopher Benton. "Structure and assembly of the sindbis virus E1 and E2 transmembrane proteins." 2006. http://www.lib.ncsu.edu/theses/available/etd-05112006-212104/unrestricted/etd.pdf.
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Hu, Yan. "Interactions between papillomavirus E1 and E2 proteins and cellular replication factors modulate each others functions." 2006. http://proquest.umi.com/pqdweb?did=1140183911&sid=5&Fmt=2&clientId=39334&RQT=309&VName=PQD.
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Thesis (Ph.D.)--State University of New York at Buffalo, 2006.Title from PDF title page (viewed on Oct. 09, 2006) Available through UMI ProQuest Digital Dissertations. Thesis adviser: Melendy, Thomas. Includes bibliographical references.
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Kajii, Takashi. "Long-term effects of prostaglandin E2 on the mineralization of a clonal osteoblastic cell line (MC3T3-E1)." Doctoral thesis, 1999. http://hdl.handle.net/2115/30170.
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共著者あり。共著者名: Kuniaki Suzuki, Masatake Yoshikawa, Tohru Imai, Akira Matsumotob and Shinji Nakamura. Elsevier Science Ltd., Takashi Kajii, Kuniaki Suzuki, Masatake Yoshikawa, Tohru Imai, Akira Matsumotob and Shinji Nakamura, Long-term effects of prostaglandin E2 on the mineralization of a clonal osteoblastic cell line (MC3T3-E1), Archives of Oral Biology, 44(3), 1999 MAR, pp.233-241. doi:10.1016/S0003-9969(98)00120-4. Journal Website: http://intl.elsevierhealth.com/journals/arob/Prostaglandin (PG) E2 is thought to be a mediator of the effect of mechanical stress on bone formation, but its effects on osteoblasts have not yet been fully described. Here, the effects of the continuous application of PGE2 and indomethacin, an inhibitor of prostaglandin G/H synthase (cyclo-oxygenase), on the proliferation, differentiation and mineralization of a clonal osteoblastic cell line, MC3T3-E1, were investigated. The cells were cultured in media with either a high (1 μg/ml) or a low (1 ng/ml) concentration of PGE2, with indomethacin (1 μg/ml) and, as a control, with neither agent. The effects of PGE2 and indomethacin were assessed quantitatively. Indomethacin and a high concentration of PGE2 increased the total protein compared to the control and low-PGE2 cultures. 7 days after confluence, alkaline phosphatase (ALP) activity within the cells and extracellular matrices increased. This increase was highest with indomethacin and lowest with a high concentration of PGE2. ALP activity also increased in the medium, but only 21 days after confluence; the effects of the agents were similar to those on the cells and matrices. The accumulation of calcium, inorganic phosphate and hydroxyproline was highest with indomethacin. PGE2 production was at its maximum when the cells were at confluence and was inhibited by indomethacin. Specific [3H]PGE2 binding to the microsomal fraction of the cell was also measured to examine the expression of the PGE2 receptor. The amount of [3H]PGE2 binding per mg of protein was highest at confluence, then decreased and again increased in the mineralizing stage. These results suggest that indomethacin increases ALP activity and the accumulation of mineralized tissue in MC3T3-E1 cells, presumably by inhibiting the production of PGE2. PGE2 could signal the suppression of mineralization as early as confluence.
Hokkaido University (北海道大学)
博士
歯学
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Mansky, Kim Carpenter. "Role of the bovine papillomavirus type E1 and E2 proteins in viral transcription and DNA replications." 1997. http://catalog.hathitrust.org/api/volumes/oclc/37900807.html.
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Thesis (Ph. D.)--University of Wisconsin--Madison, 1997.Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 178-198).
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Pelzer, Christiane [Verfasser]. "Characterization of novel E1 and E2 enzymes and their role in ubiquitin and FAT10 conjugation / vorgelegt von Christiane Pelzer." 2009. http://d-nb.info/1011541882/34.
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Franken, Tobias [Verfasser]. "Generierung, Charakterisierung und funktionaler Assay von Antikörpern und Antikörperfragmenten gegen die HCV-Strukturproteine core, E1 und E2 / vorgelegt von Tobias Franken." 2008. http://d-nb.info/990706370/34.
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Clower, Randolph Vincent. "The papillomavirus E1 helicase and E2 protein bind to and stimulate the enzymatic functions of human topoisomerase I and DNA polymerase delta." 2006. http://proquest.umi.com/pqdweb?did=1184160381&sid=7&Fmt=2&clientId=39334&RQT=309&VName=PQD.
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Thesis (Ph.D.)--State University of New York at Buffalo, 2006.Title from PDF title page (viewed on Mar. 20, 2007) Available through UMI ProQuest Digital Dissertations. Thesis adviser: Melendy, Thomas E. Includes bibliographical references.
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Schmidt, Matthias [Verfasser]. "Herstellung und Charakterisierung von virusähnlichen Partikeln auf der Basis von Fusionsproteinen aus HBV-Coreprotein und HCV-Oberflächenproteinen E1 bzw. E2 / vorgelegt von Matthias Schmidt." 2008. http://d-nb.info/998515760/34.
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Castelão, Cindy Duarte 1990. "Papel dos mecanismos envolvendo esteróides sexuais (E1 e E2), sua variação genética e biomarcadores circulantes na etiopatogenia de tumores ginecológicos (leiomiomas e cancro do colo do útero)." Master's thesis, 2013. http://hdl.handle.net/10451/10028.
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Tese de mestrado. Biologia (Biologia Molecular e Genética). Universidade de Lisboa, Faculdade de Ciências, 2013Os leiomiomas são neoplasias benignas que se formam a partir das células do músculo liso. Estes são os tumores mais comuns do aparelho reprodutor feminino. O cancro do colo do útero, cujo agente patogénico é o HPV, é o segundo cancro mais frequente e a segunda maior causa de morte nas mulheres em todo o mundo. Um dos factores de risco para o desenvolvimento de neoplasias em tecidos sensíveis a hormonas é a exposição, excessiva e cumulativa a estrogénios. Um factor importante da toxicologia dos estrogénios, para além da estimulação da proliferação de células epiteliais, é o seu metabolismo oxidativo. Assim, estudou-se polimorfismos funcionais nas seguintes enzimas: o CYP1A1 (rs4646903), a COMT (rs 4680), a MPO (rs2333227). Estudou-se igualmente a actividade da RTM e a concentração de estradiol circulante. Neste estudo observou-se que o alelo C, responsável pela maior actividade de CYP1A1, é um factor de risco em ambas as patologias. O alelo A, que concede menor actividade à enzima COMT, apresenta-se como um factor de risco para as patologias em estudo. Observámos que o genótipo GA da enzima MPO revelou-se um factor de risco em ambas as patologias – isto poderá indicar que esta é importante na eliminação do HPV; ou poderá ser um indício de que quantidades excessivas de ROS são prejudiciais às células transformadas. A RTM não demonstrou resultados estatisticamente significativos. Os níveis de estradiol circulante foram superiores nas populações patológicas corroborando a teoria que esta hormona é tumorigénica, quer directa quer indirectamente.A maior concentração de estradiol na população de leiomiomas pode indicar uma maior dependência hormonal deste. O facto dos resultados obtidos serem idênticos nos leiomiomas e cancro do colo do útero é interessante: são dois tumores diferentes, não só na sua etiologia, mas também na sua fisiologia; no entanto, os riscos não se distinguiram, indicando aparência em termos de susceptibilidade.
Leiomyomas are benign neoplasms that arise from smooth muscle cells. These are the most common tumors of the feminine reproductive system. HPV infection may lead to cervical cancer; this is the second most frequent cancer and the second biggest cause of death in women worldwide. One risk factor for the development of neoplasms in hormone-sensitive tissue in women is the excessive and cumulative exposure to estrogens. Estrogen toxicology can be explained by its hability to promote proliferation of epithelial cells, but also because of its oxidative metabolism. So, we studied the functional polymorphisms in the following enzymes: CYP1A1 (rs4646903), COMT (rs 4680), and MPO (rs2333227). We also studied the activity of RTM, and the concentration of estradiol. In this study we observed that the allele C, responsible for a more active CYP1A1, it’s a risk factor in both pathologies. The allele A, that codifies a lower activity COMT, also presents itself as a risk factor for both leiomyomas and cervical cancer. The GA genotype in the MPO enzyme revealed itself as a risk factor for both pathologies – which may indicate that MPO is important in the elimination of HPV from the organism; or that excessive amounts of ROS could lead to apoptosis of tumorigenic cells. The analysis of RTM activity demonstrated no significant statistical values. The levels of estradiol in blood were superior on the studied populations, supporting the theory that this hormone is tumorigenic, either acting directly or through its metabolism. The results obtained through this study demonstrate that both leiomyomas and cervical cancer show similar susceptibilities, even though they are very different from each other: not only in their etiology but also in their physiology.
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Masavuli, Makutiro Ghislain. "Novel DNA Vaccine Formulations Against Hepatitis C Virus." Thesis, 2018. http://hdl.handle.net/2440/127111.
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No vaccines are available for hepatitis C virus (HCV) which infects over 71 million people worldwide. Current therapeutic options are very expensive and as a consequence, only around 1% of those diagnosed with Hepatitis C receive treatment each year. Induction of neutralizing antibody (NAb) by vaccination will be important for the successful prevention of HCV infection. HCV envelope glycoproteins E1 and E2 are required for virus entry into host cells making these proteins attractive targets to prevent virus infection. DNA-based vaccines are appealing candidates for novel vaccine development because they are not infectious and are cost-effective to manufacture on a global scale. However, despite being licensed for veterinary use, DNA vaccine have not been highly immunogenic in large animals. Virus-like particles (VLP), on the other hand, which resemble native viruses but are non-infectious because they lack viral genetic materials, have provided highly encouraging results in clinical trials. In this thesis the immunogenicity of VLPs and the ease of production of plasmid DNA were combined by designing DNA vaccines encoding VLPs consisting of HCV-core, E1 and E2 proteins (which can self-assemble into VLPs following expression). This vaccine also encoded a cytolytic gene perforin (PRF) to cause cell death and ensure the release of the VLPs from vaccine-targeted cells along with damage associated molecular patterns (DAMPs), which act as natural adjuvants. Bicistronic DNA vaccine constructs encoding HCV structural proteins and PRF were successfully generated and validated. Vaccination with the DNA construct encoding CoreE1E2-PRF generated higher adaptive immune responses in mice than vaccination with a construct unable to induce cell death, therefore confirming an adjuvant effect by PRF. However, these responses were weak compared to those reported in the literature. Antigen oligomerisation has been shown to improve vaccine immunogenicity. To improve the immunogenicity of a DNA vaccine encoding HCV-E1E2, a novel strategy that incorporates E1 and E2 into heptamers by fusion with the oligomerisation domain of a chimeric C4 binding protein (namely IMX3133 or IMX313P) was used. As the adjuvanticity of IMX313 or IMX313P requires efficient secretion of the oligomerised protein, the leader sequence of the tissue plasminogen activator (tPA) was introduced upstream of the E1 or E2 proteins (tE1 or tE2) from which the transmembrane domains were removed. The use of tE1 and tE2 proteins as separate immunogens or as a single tE1tE2 polyprotein when fused to IMX313 or IMX313P was assessed in vaccination studies in Balb/c mice using prime-boost intra-dermal DNA immunisations. Vaccination with the DNA construct encoding tE1/tE2 fused to IMX313 or IMX313P resulted in increased antibody and cell mediated immune (CMI) responses compared to the same dose of DNA without IMX313 or IMX313P, while fusion of tE1/tE2 to IMX313P resulted in the highest immune responses. DNA prime/E1E2 protein boost or DNA prime/HCV-VLP boost approaches were then used to further improve the immunogenicity of tE1/tE2-IMX313P DNA constructs. Boosting with E1E2 proteins improved overall antibody responses, compared to boosting with HCV-VLPs or plasmid DNA. However, experiments to examine the neutralisation of binding of labelled VLPs showed that the E1E2 or VLP boost did not improve the neutralising potency of these antibodies or the CMI responses. This thesis demonstrated that expression of heptamerised and secreted tE1/tE2 from DNA vaccine constructs significantly improved the antibody and CMI responses to HCV E1 and E2 proteins. Most importantly, the antibody induced by these constructs possess neutralising properties.Thesis (Ph.D.) -- University of Adelaide, Adelaide Medical School, 2018
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Nevzorova, Yulia [Verfasser]. "Cell cycle regulation in the liver: differential functions of E-type cyclins E1 and E2 for G1/S-phase transition and endoreplication in mice / vorgelegt von Yulia Nevzorova." 2009. http://d-nb.info/1004859481/34.
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