A proteína ribossomal L10 (RP L10) é uma forte candidata a ser incluída na classe de proteínas supressoras de tumor. Também denominada QM, a proteína em questão é conhecida por participar da ligação das subunidades ribossomais 60S e 40S e da tradução de mRNAs. Possui massa molecular entre 24 a 26 kDa e ponto isoelétrico (pI) 10,5. A seqüência da proteína QM é bastante conservada em mamíferos, plantas, invertebrados, insetos e leveduras indicando que esta possui funções críticas na célula. Com função supressora de tumor, a proteína RP L10 foi estudada em linhagens de tumor de Wilm (WT-1) e em células tumorais de estômago, nas quais se observou uma diminuição na quantidade de seu mRNA. Mais recentemente a RP L10 foi encontrada em baixas quantidades nos estágios iniciais de adenoma de próstata e com uma mutação em câncer de ovário, indicando uma participação no desenvolvimento destas doenças. Como proteína, já foi descrito que esta interage com as proteínas c-Jun e c-Yes, inibindo a ação ativadora de fatores de crescimento e divisão celular. Este trabalho tem um papel importante no estabelecimento da expressão desta proteína solúvel, para estudos posteriores que tenham como objetivo avaliar a ação de regiões específicas que atuam na ligação das subunidades ribossomais 60S e 40S e tradução, bem como nas regiões que se ligam a proto-oncogenes. O cDNA para proteína QM foi amplificado por PCR e clonado no vetor de expressão periplásmica p3SN8. A proteína QM foi expressa em E.coli BL21 (DE3) no citoplasma e periplasma bacteriano e na melhor condição, a expressão de QM de bactérias transformadas pelo plasmídeo recombinante p1813_QM em 25°C ou 30°C, a proteína foi obtida solúvel e com quantidad es muito pequenas de contaminantes. Os ensaios de estrutura secundária demonstraram que a proteína QM tem predominância de a-hélice, mas quando do seu desenovelmento, essa condição muda e a proteína passa a ter característica de folhas β. / The ribosomal protein L10 (RP L10) is a strong candidate to be included in the class of tumor suppressor proteins. This protein, also denominated as QM, is known to participate in the binding of ribosomal subunits 60S and 40S and the translation of mRNAs. It has a molecular weight that varies between 24 and 26 kDa and an isoelectric point of (pI) 10.5. The sequence of the protein QM is highly conserved in mammals, plants, invertebrates, insects and yeast which indicates its critical functions in a cell. As a tumor suppressor, RP L10 has been studied in strains of Wilm\'s tumor (WT-1) and tumor cells in the stomach, where was observed a decrease in the amount of its mRNA. More recently, the RP L10 was found in low amounts in the early stages of prostate adenoma and showed some mutation in ovarian cancer, what indicates its role as a suppressor protein in the development of these diseases. It has also been described that this protein interacts with c-Jun and c-Yes inhibiting growth factors and consequently, cell division. This work has an important role on the establishment of soluble expression of QM to give base information for further studies on expression that aim to evaluate the specific regions where it acts binding the 60S and 40S ribossomal subunits and translation, as well as its binding to proto-oncogenes. The cDNA for QM protein was amplified by PCR and cloned into periplasmic expression vector p3SN8. The QM protein was expressed in E. coli BL21 (DE3) in the region of cytoplasm and periplasm, the best condition was obtained from the expression of the recombinant plasmid QM p1813_QM at 25°C or 30°C, the soluble protein was obtained with small amounts of contaminants. The assays of secondary structure showed that the QM protein is predominantly alpha-helix, but when it loses the folding, this condition changes and the protein is replaced by β- sheet feature.

A proteína ribossomal L10 (RP L10) é uma forte candidata a ser incluída na classe de proteínas supressoras de tumor. Também denominada QM, a proteína em questão é conhecida por participar da ligação das subunidades ribossomais 60S e 40S e da tradução de mRNAs. Possui massa molecular entre 24 a 26 kDa e ponto isoelétrico (pI) 10,5. A seqüência da proteína QM é bastante conservada em mamíferos, plantas, invertebrados, insetos e leveduras indicando que esta possui funções críticas na célula. Com função supressora de tumor, a proteína RP L10 foi estudada em linhagens de tumor de Wilm (WT-1) e em células tumorais de estômago, nas quais se observou uma diminuição na quantidade de seu mRNA. Mais recentemente a RP L10 foi encontrada em baixas quantidades nos estágios iniciais de adenoma de próstata e com uma mutação em câncer de ovário, indicando uma participação no desenvolvimento destas doenças. Como proteína, já foi descrito que esta interage com as proteínas c-Jun e c-Yes, inibindo a ação ativadora de fatores de crescimento e divisão celular. Este trabalho tem um papel importante no estabelecimento da expressão desta proteína solúvel, para estudos posteriores que tenham como objetivo avaliar a ação de regiões específicas que atuam na ligação das subunidades ribossomais 60S e 40S e tradução, bem como nas regiões que se ligam a proto-oncogenes. O cDNA para proteína QM foi amplificado por PCR e clonado no vetor de expressão periplásmica p3SN8. A proteína QM foi expressa em E.coli BL21 (DE3) no citoplasma e periplasma bacteriano e na melhor condição, a expressão de QM de bactérias transformadas pelo plasmídeo recombinante p1813_QM em 25°C ou 30°C, a proteína foi obtida solúvel e com quantidad es muito pequenas de contaminantes. Os ensaios de estrutura secundária demonstraram que a proteína QM tem predominância de a-hélice, mas quando do seu desenovelmento, essa condição muda e a proteína passa a ter característica de folhas β. / The ribosomal protein L10 (RP L10) is a strong candidate to be included in the class of tumor suppressor proteins. This protein, also denominated as QM, is known to participate in the binding of ribosomal subunits 60S and 40S and the translation of mRNAs. It has a molecular weight that varies between 24 and 26 kDa and an isoelectric point of (pI) 10.5. The sequence of the protein QM is highly conserved in mammals, plants, invertebrates, insects and yeast which indicates its critical functions in a cell. As a tumor suppressor, RP L10 has been studied in strains of Wilm\'s tumor (WT-1) and tumor cells in the stomach, where was observed a decrease in the amount of its mRNA. More recently, the RP L10 was found in low amounts in the early stages of prostate adenoma and showed some mutation in ovarian cancer, what indicates its role as a suppressor protein in the development of these diseases. It has also been described that this protein interacts with c-Jun and c-Yes inhibiting growth factors and consequently, cell division. This work has an important role on the establishment of soluble expression of QM to give base information for further studies on expression that aim to evaluate the specific regions where it acts binding the 60S and 40S ribossomal subunits and translation, as well as its binding to proto-oncogenes. The cDNA for QM protein was amplified by PCR and cloned into periplasmic expression vector p3SN8. The QM protein was expressed in E. coli BL21 (DE3) in the region of cytoplasm and periplasm, the best condition was obtained from the expression of the recombinant plasmid QM p1813_QM at 25°C or 30°C, the soluble protein was obtained with small amounts of contaminants. The assays of secondary structure showed that the QM protein is predominantly alpha-helix, but when it loses the folding, this condition changes and the protein is replaced by β- sheet feature.

Nous avons étudié l'anisotropie magnétique perpendiculaire résultant de la mise en ordre chimique de type L10, dans des couches minces d'alliage FePd élaborées en Epitaxie par Jets Moléculaires. Différentes procédures d'élaboration ont été mises en oeuvre : - la codéposition à température ambiante, éventuellement suivie d'un recuit ; - la codéposition à 350°C ; - le dépôt alterné de couches atomiques Fe et Pd, contrôlé par le temps de dépôt ou par les oscillations RHEED. La structure des alliages a été étudiée par Microscopie Electronique en Transmission. Nous avons caractérisé l'ordre à longue distance (OLD) par diffraction des rayons X, et l'ordre à courte distance directionnel (OCDD) par spectroscopie EXAFS. L'anisotropie magnétique uniaxiale a été évaluée à partir de mesures de magnétométrie (VSM). Nous montrons que l'OLD et l'OCDD, de même que l'anisotropie magnétique, dépendent fortement des conditions de dépôt. Le degré d'ordre chimique le plus élevé est obtenu par la codéposition de l'alliage à 350°C : dans ce cas, l'aimantation est orientée suivant la direction perpendiculaire au plan des couches minces et l'étude par Microscopie à Force Magnétique révèle la présence de domaines magnétiques, dont la taille latérale est de l'ordre de quelques dizaines de nanomètres. L'anisotropie magnétique résultant du dépôt alterné de couches atomiques est plus faible : pour les faibles épaisseurs, l'aimantation est dans le plan de la couche et au delà d'une épaisseur critique, elle sort du plan, faisant apparaître une configuration en rubans. Nous avons interprété, par des modèles analytiques de micromagnétisme, l'évolution de la susceptibilité en champ perpendiculaire, ainsi que celle de la taille des domaines et des rubans, en fonction de l'épaisseur des couches minces. Nous avons également réalisé des expériences de spectroscopie Mössbauer et spectroscopie Kerr polaire : nous montrons que ces deux signaux sont très sensibles au degré d'ordre des alliages.

No description available.

L'objectif de ce travail de thèse est de préparer des fils de CoPt et FePt par dépôt électrochimique dans des membranes d'alumine nanoporeuse. Le bain électrochimique que nous avons utilisé comprend seulement deux sels (des chlorures), un pour le cobalt (CoCl2, 6H2O) et un pour le platine (K2PtCl6). Nous avons réussi à réaliser des réseaux de nanofils de CoPt ayant des diamètres d'environ 70-80 nm et dont la coercitivité atteint 1.1 Tesla à température ambiante. Ces matériaux magnétiquement durs présentent la phase quadratique L10 obtenue après un recuit à 700 °C de la phase cubique déposée.<br />Nous avons étudié les propriétés structurales et magnétiques d'échantillons préparés dans les deux types de membranes que nous avons utilisées: des membranes faites au laboratoire par anodisation d'une couche d'aluminium et des membranes commerciales. Les mesures magnétiques effectuées parallèlement et perpendiculairement à l'axe des fils montrent un comportement parfaitement isotrope. La qualité des deux types de réseau de pores utilisés ne semble pas avoir d'influence sur les propriétés magnétiques et en particulier sur le champ coercitif des échantillons étudiés. Les paramètres (température, durée) du recuit sont les paramètres essentiels pour obtenir la transformation la plus complète possible de la phase cubique déposée en la phase quadratique ordonnée L10 .<br />La préparation de nanofils de FePt s'est révélée plus compliquée du fait de la facilité du fer à former des oxydes. Nous avons tout de même réussi à préparer des réseaux de nanofils de FePt de 55~nm de diamètre avec une coercitivité de 1.1 Tesla mais non homogènes en composition.

No description available.

The large magneto-crystalline anisotropy energy of the L10 phase has pushed the interest to the FePt nanoparticles to get smallest possible not superparamagnetic particles for magnetic data storage media. The DC magnetron sputtering process, in an inert gas atmosphere and subsequently ejection into high vacuum via differential pumping in addition with a newly constructed light furnace, allows us to have a predeposition annealing of FePt nanoparticles. The advantage compared to wet chemical process route is, that we can prevent the growing of particles on a substrate. In order to determine the experimentally hardly accessible temperature of the particles, the thermal history of the particles is rather calculated from the interaction with the light field along the flight path through the light furnace used for the in-flight annealing. The results obtained for the particle temperature are corroborated by experimental findings on the sintering of agglomerated particles and change in magnetic properties due to heating over the L10 stability temperature. The experiments reveal that the effect of the thermal treatment on both the structural and magnetic properties of the FePt nanoparticles strongly depends on the particles’ crystal structure. The magnetic behavior shows a size depending effective uniaxial magnetic anisotropy constant. This behavior is strongly correlated to the structure of the 5 nm to 8 nm L10 FePt particle.

Made available in DSpace on 2015-03-26T13:42:37Z (GMT). No. of bitstreams: 1 texto completo.pdf: 1636951 bytes, checksum: 5c4c3aea8b56b9b0913479b27b012cea (MD5) Previous issue date: 2007-08-01 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Proteins of the family of LRR-RLKs (receptor-like-kinases with leucine-rich repeats) have a conceptual relevance in signaling events but in plants information regarding function is limited to a few members of this family. The receptors NIK1, NIK2 and NIK3 of Arabidopsis thaliana belong to the sub-family LRRII-RLK and were initially identified by their capacity to interact with the geminivirus NSP protein. In response to an unknown stimulus, NSP-interacting kinases (NIKs) are activated after the formation of dimmers and intermolecular autophosphorylation. The inhibition of autophosphorylation of NIK by NSP and the activation of NIK genes increase the susceptibility to viral infection, suggesting that this protein is involved in a defense pathway against geminivirus infection. The downstream components of this pathway, mediated by the protein NIK, have yet to be identified. In the present study, the biochemical and functional characterization of two ribosomal proteins, L10 and L18 were described, these being capable of interacting with the protein NIK via the yeast two-hybrid system. In vitro studies demonstrated that the protein NIK is capable of phosphorylating the protein L10, but not L18. Of the members of the LRRIIRLK family, the protein NIK2 phosphorylates L10, while NIK3 presents a low capacity for phosphorylation of the substrate. However, the development protein SERK1 does not use L10 as a substrate. Assays of transient expression in tobacco plants, revealed that the L18 protein is located in the cytoplasm, as well as around the nucleus and the nucleoli of some cells. In turn, in 97% of the cells, L10 was localized only in the cytoplasm, although it was also found in the nucleus, in approximately 3% of the observed cells. The transient expression of NIK1 and NIK2 redirects the L10 protein to the nucleus in approximately 30% of the cells. In contrast, NIK3 does not relocalize the L10 protein to the nucleus, and L18 does not change its localization in the presence of the NIKs. In plants infected with TGMV, a change only in the cytoplasmic localization of L10 was observed, accumulating in points of the cytoplasm when not co-localized with NIK. To prove genetically the interactions of L10-NIK and L18-NIK, null alleles for the genes L10 and L18 de Arabidopsis, containing T-DNA insertion, were obtained and inoculated with CaLCuV. The inactivation of the genes L10 and L18 restored the elevated susceptibility phenotype of nik1 and the knockout plants, principally l10, presented severe symptoms and high rates of infection when compared with the wild columbia plants. The results of this work are consistent with a model that places the ribosomal proteins L10 and L18 as functional components of the defense signaling pathway mediated by the protein NIK, L10 being a component immediately downstream of the transmembrane receptor. / As proteínas da família das LRR-RLKs (receptor-like-quinases com repetições ricas em leucina) possuem uma relevância conceitual em eventos de sinalização mas, em plantas, a informação funcional ainda é restrita a alguns membros desta família. Os receptores NIK1, NIK2 e NIK3 de Arabidopsis thaliana pertecem à sub-familia LRRII-RLK e foram inicialmente identificados pela sua capacidade de interagir com a proteína NSP de geminivírus. Em resposta a um estímulo desconhecido, NSP-interacting kinases (NIKs) são ativadas após a formação de dímeros e autofosforilação intermolecular. A inibição da autofosforilação de NIK por NSP e a inativação de genes NIKs aumenta a suscetibilidade à infecção viral, sugerindo que esta proteína estaria envolvida em uma via de defesa contra a infecção por geminivírus. Os componentes downstream dessa via de sinalização, mediada pela proteína NIK, ainda não foram identificados. No presente estudo foi descrita a caracterização bioquímica e funcional de duas proteínas ribossomais, L10 e L18, as quais foram capazes de interagir com a proteína NIK através do sistema de duplo híbrido de leveduras. Estudos in vitro demonstraram que a proteína NIK é capaz de fosforilar a proteína L10, mas não L18. Entre os membros da família LRRII-RLK, a proteína NIK2 fosforila L10, enquanto NIK3 apresenta uma baixa capacidade de fosforilação do substrato. No entanto, a proteína de desenvolvimento SERK1 não utiliza L10 como substrato. Ensaios de expressão transiente, em plantas de tabaco, revelaram que a proteína L18 está localizada no citoplasma, bem como ao redor do núcleo e no nucléolo de algumas células. Por sua vez, em 97% das células, L10 foi localizada apenas no citoplasma, embora tenha sido encontrada no núcleo, em aproximadamente 3% das células observadas. A expressão transiente de NIK1 e NIK2 redireciona a proteína L10 para o núcleo em aproximadamente 30% das células. Em contraste, NIK3 não relocaliza a proteína L10 para o núcleo, e L18 não muda sua localização na presença das NIKs. Em plantas infectadas com TGMV, observou-se mudança apenas na localização citoplasmática de L10, acumulando-se em pontos do citoplasma quando não colocalizada com NIK. Para se comprovar geneticamente as interações de L10-NIK e L18- NIK, alelos nulos para os genes L10 e L18 de Arabidopsis, contendo inserção de T-DNA, foram obtidos e inoculados com o CaLCuV. A inativação dos genes L10 e L18 recapitulou o fenótipo de suscetibilidade aumentada de nik1 e as plantas knockout, principalmente l10, apresentaram sintomas severos e taxa de infecção alta quanto comparados com as plantas selvagens columbia. Os resultados deste trabalho são consistentes com um modelo que posiciona as proteínas ribossomais L10 e L18 como componentes funcionais da via de sinalização de defesa mediada pela proteína NIK, sendo L10 um componente imediatamente downstream ao receptor transmembrana.

We study the effects of the recent economic crisis on firms' bidding behavior and markups in sealed bid auctions. Using data from Austrian construction procurements, we estimate bidders' construction costs within a private value auction model. We find that markups of all bids submitted decrease by 1.5 percentage points in the recent economic crisis, markups of winning bids decrease by 3.3 percentage points. We also find that without the government stimulus package this decrease would have been larger. These two pieces of evidence point to pro-cyclical markups. (authors' abstract)

Highly ordered magnetic materials with high perpendicular magnetic anisotropy (PMA), such as the L10 ordered FePt, and new recording technologies, such as bit patterned media (BPM), have been proposed as solutions to the media trilemma problem and provide promising strategies towards future high-density magnetic data storage media. L10 ordered FePt thin films can provide the necessary high PMA. However, the ordering of this material perpendicular to the plane of the films remains challenging since high-temperature and time-consuming processes are required. In this work, a remote plasma sputtering system has been used for the investigation of FePt thin films in order to understand if the greater control of process parameters offered by this system can lead to enhanced ordering in L10 FePt thin films at low temperatures compared with conventional dc magnetron approaches. More specifically, the effect of the different substrate temperatures and the target bias voltages on the ordering, the microstructure and the magnetic properties of FePt thin films was investigated. Highly ordered FePt thin films were successfully fabricated after post-annealing processes and were patterned into arrays of FePt islands. This patterning process was carried out with e-beam lithography and ion milling. Initial MFM measurements of these islands showed their single-domain structure for all the island sizes, which indicated the high PMA of the FePt. Magnetometry measurements were also carried out with a novel polar magneto-optical Kerr effect (MOKE) system which was designed and built during this project. This system has unique capabilities which are: a) the application of uniform magnetic field up to 2 Tesla, b) the rotation of the field to an arbitrary angle and c) the use of lasers of four different wavelengths. The combination of these abilities enabled measurements on ordered FePt thin films and patterned media which can pave the way for further highly sensitive measurements on magnetic thin films and nanostructures.