Dissertations / Theses on the topic 'Baculum'

Les hydrogenases sont des enzymes qui catalysent la production ou la consommation de l'hydrogene moleculaire selon la reaction suivante : h#2 2h#+ +2e. L'hydrogenase de desulfovibrio gigas existe sous differents etats redox, appeles etats inactif, pre-actif (formes oxydees) et actif (forme reduite). La structure de l'hydrogenase de d. Gigas dans son etat oxyde, a une resolution de 2,85a, a montre que la proteine contenait deux agregats 4fe4s et un agregat 3fe4s, situes dans la petite sous-unite. Le site actif est enfoui dans la grande sous-unite et contient un atome de nickel, ainsi qu'un autre site metallique a proximite du nickel, probablement un fer et appele atome x. Une experience de dispersion anomale au seuil d'absorption du fer a montre pour la premiere fois que le site actif etait compose d'un atome de nickel et d'un atome de fer a une distance de 3a. Les deux metaux sont coordonnes par quatre cysteines dont deux sont pontantes entre les deux metaux ; l'atome de fer possede egalement trois ligands de nature inconnue et modelises comme trois molecules d'eau. L'analyse de la structure de la forme oxydee obtenue a 2,54a de resolution sur une autre forme cristalline a montre la presence d'un ligand pontant entre les deux metaux, modelise comme une espece oxygenee. D'autre part, en parallele avec des etudes infrarouges sur l'hydrogenase de chromatium vinosum, les trois ligands du fer ont ete attribues a des especes diatomiques du type co/cn. Une etude des differents etats redox est necessaire afin de mieux comprendre le mecanisme catalytique. L'installation d'une boite a gants ainsi que la mise au point de techniques de reduction et de congelation des cristaux ont donc ete realisees. La structure de l'hydrogenase a ni-fe-se de desulfomicrobium baculatum dans son etat reduit a ete determinee a une resolution de 2,15a par la methode du remplacement moleculaire. D'autre part, nous avons egalement obtenu la structure de l'hydrogenase de d. Gigas sous sa forme reduite a une resolution de 2,7a. Une comparaison des sites actifs des deux hydrogenases dans les etats oxyde et reduit fait apparaitre un certain nombre de differences. Sur la base des differentes structures, nous pouvons proposer un mecanisme d'activation.

Orientador : Yoko Bomura Rosato
Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: Bacillus thuringiensis subsp. Israelensis é uma bactéria entomopatogênica que produz cristais protéicos durante o processo de esporulação. Com o objetivo de se determinar a atividade biológica das proteínas constituintes do cristal foi feita a clonagem molecular dos genes da toxina. A estratégia de clonagem consistiu-se na inserção de fragmentos de DNA plasmidial de B. thuringiensis subsp israelensis no plasmídio pUC8, resultando em plasmídios híbridos. Foram obtidos 8 clones recombinantes com atividade contra larvas de ª fluviatilis e hemolíticos para eritrócitos de carneiros e um clone recombinante com atividade larvicida e não hemolítico. Após análise imunológica com anticorpos produzidos contra as principais frações protéicas de B. thuringiensis subsp israelensis, verificou-se a produção dos peptídeos de 28, 33 e 38 kD nos clones recombinantes larvicidas e hemolíticos e a ausência do peptídeo de 33 kD no clone não hemolítico
Abstract: Bacillus thuringiensis subsp. Isralensis is a bacteria entomopathogenic that produce proteins crystals during the process of sporulation. The molecular cloning of the toxin genes was carried out to determine the biological activities of the crystal proteins. Fragments of DNA plasmids of Bacillus thuringiensis subsp. Israelensis coli were inserted in the vector pUC8 and cells of Escherichia coli were transformed with the hybrid plasmids. Eight of the recombinants obtained were active against larvae of A.fluviatilis and hemolytic for sheep red blood cells, while one of the recombinants showed larvicidal activity but did not show hemolytic activity. Antibodies were produced against the major proteins of Bacillus thuringiensis subsp. israelensis crystal and the immunodection test showed the presence of three types of proteins with 28, 33 and 38 kD, respectively. While the clones with IarvicidaI and hemolytic activity contained all three types of proteins, the one with no hemolytic activity lacked the 33 kD protein
Mestrado
Genetica
Mestre em Ciências Biológicas

Arrabidaea chica Verlot (Bignoniaceae), conhecida como crajiru, fornece pigmentos vermelhos utilizados pelos índios no Brasil como corante e agente cicatrizante. Este estudo visou otimizar a extração de compostos fenólicos de A. chica e avaliar sua atividade farmacológica. Extratos de A. chica foram obtidos através de tratamento com xilanases de Bacillus pumilus previamente à extração. Os ensaios foram monitorados por CLAE e ESI-MS. O tratamento enzimático forneceu extratos enriquecidos em antocianidinas. Extratos sem tratamento enzimático apresentaram maior teor de antocianosídeos. O estudo farmacológico demonstrou que as atividades anticâncer e antioxidante in vitro estão diretamente relacionadas ao maior teor de agliconas. O ensaio in vitro de indução de crescimento de fibroblastos indicou que o maior teor da aglicona carajurina é inversamente proporcional à ação cicatrizante. Portanto, foi desenvolvida uma metodologia inovadora, através de processos biotecnológicos, para extração de antocianidinas que apresentam propriedades corantes e terapêuticas.
Arrabidaea chica Verlot (Bignoniaceae), known as crajiru, produces red pigments used by Brazilian indians as dye and as healing agent. This study has aimed the optimization of phenolic compounds extraction from A. chica and to evaluate its pharmacological activities. Extracts from A. chica were obtained through treatment with xylanases from Bacillus pumilus before the extraction. The assays were monitored by HPLC and ESI/MS-MS. The enzymatic treatment has produced more concentrated in anthocyanidins extracts. Those obtained without enzymatic treatment have presented higher glycosilated anthocyanins content. The pharmacologic study has demonstrated that the antitumoral and the antioxydant in vitro properties for A. chica are directly related to the higher contents of aglycones. In vitro assay for fibroblasts growth induction has demonstrated that a higher content of carajurin is inversely proportional to the healing action. In conclusion, a novel approach has been developed, through biotechnological process, aiming the extraction of anthocyanidins presenting dye and therapeutic properties.

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Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico
Societal concerns about environmental sustainability has lead to the development of ecologically-friendly alternatives to chemical insecticides for crop protection. One such alternative is biological pest control. In particular, baculoviruses are well suited as insect biopesticides due to their narrow host specificity and relative ease of propagation. In Brazil, the baculovirus Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV) is the main biological control agent employed for the soybean pest, Anticarsia gemmatalis. This baculovirus biopesticide is currently produced using caterpillars, but increasing market demand for the product has encouraged the development of an in vitro manufacturing process, which can be scaled up to much higher virus productivities. In this study, three wild-type AgMNPV isolates (AgMNPV-2D, AgMNPV-MP2 and AgMNPV-MP5) and a recombinant form (vAgEGT-LacZ) were characterised in terms of occlusion body (OB) production and infection kinetics, to enable future optimisation of the in vitro production process. These viruses were propagated using a Spodoptera frugiperda (IPLB-SF21) insect cell line grown in shaker-flask batch cultures. Among the virus isolates tested, AgMNPV-MP5 was found to be the best producer, yielding (5.3?0.85)x108 OB/mL after 8 days post infection. The characterisation of vAgEGT-LacZ propagation in suspension cell cultures has not been previously reported in the literature; hence it became the main focus for this thesis. In particular, it was carried out a study on the effect of the multiplicity of infection (MOI) on OB production. Five successive batches were performed getting a final production (8.9?1.42)x1014 occlusion bodies, considering that production is related for a bioreactor with final volume of 10m3. A low MOI associated with a fed-batch process for vAgEGT-LacZ production was found to support a 3-fold higher OB yield when compared to the default batch process (1.8x107 and 5.3x107 OB/mL, respectively). This yield is competitive with regards to the production process.
A preocupa??o da sociedade com o meio ambiente tem levado a buscar alternativas de substitui??o dos inseticidas qu?micos por outros produtos menos agressivos ao homem e ao ambiente. Assim, a utiliza??o de controle biol?gico contra pragas ? altamente desej?vel, pois reduz os riscos ambientais e p?blicos da utiliza??o de produtos qu?micos convencionais. Em particular, os v?rus do tipo baculov?rus s?o uma grande alternativa devido ? especificidade oferecida aos seus hospedeiros e a sua forma de propaga??o. No Brasil, o baculov?rus Anticarsia gemmatalis multiple nucleopolyhedrovirus (AgMNPV) ? o principal agente de controle biol?gico da praga da soja Anticarsia gemmatalis. A crescente demanda deste bioinseticida tem estimulado o interesse no desenvolvimento de processos com base na produ??o in vitro de baculov?rus. Deste modo, poder? aumentar a oferta de v?rus, suprindo a necessidade do mercado deste bioinseticida para o controle de A. gemmatalis. Neste trabalho, estrat?gias de produ??o in vitro do baculov?rus selvagem AgMNPV e do seu recombinante vAgEGT?-LacZ, como influ?ncia do isolado, da cin?tica e do modo de opera??o, foram analisadas como alternativa para futura amplia??o de escala na produ??o deste bioinseticida viral. A produ??o em batelada de tr?s isolados selvagens do baculov?rus AgMNPV (AgMNPV-2D, AgMNPV-MP2 e AgMNPV-MP5) utilizando o cultivo em shaker, foi realizada com a finalidade de selecionar o melhor produtor de corpos de oclus?o a partir das infec??es em c?lulas de inseto Spodoptera frugiperda, linhagem IPLB-SF21 para avalia??o comparativa com recombinante vAgEGT?-LacZ. A sele??o identificou o isolado AgMNPVMP5 como melhor produtor de corpos de oclus?o de (5,30?0,85) x108 OB/mL em 8 dias de infec??o. A produ??o in vitro do vAgEGT?-LacZ foi foco principal deste trabalho, pois n?o h?, na literatura, a produ??o deste recombinante em sistemas de cultivo em suspens?o. Foi realizado estudo da multiplicidade de infec??o para identificar a quantidade de in?culo viral para o cultivo. A partir da?, foram realizadas cinco bateladas sucessivas obtendo-se (8,9?1,42)x1014 corpos de oclus?o para um volume final de 10m? de suspens?o de c?lulas infectadas. O aumento da produ??o de corpos de oclus?o obtidos a partir do vAgEGT?-LacZ foi analisado utilizando a estrat?gia de cultivo em batelada alimentada utilizando baixa multiplicidade de infec??o. Esta estrat?gia permitiu aumento de tr?s vezes na produ??o de corpos de oclus?o quando comparada ? produ??o em cultivos em batelada (5,3x107 e 1,8x107 OB/mL, respectivamente). E ainda, o baculov?rus recombinante vAgEGT?-LacZ mostrou-se competitivo em rela??o ao baculov?rus selvagem AgMNPV-MP5

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Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico
Among the pests that attack corn crop in Brazil, there is Spodoptera frugiperda (JE Smith, 1797) (Lepidoptera: Noctuidae), known as fall armyworm, which is the major corn pest. Due to genetic instability during serial passage of baculoviruses in insect cell culture, the viral bioinseticides in vitro production development is the greatest challenge for mass production of this bioproduct. Successive passages of virus using extracellular viruses (BVs), necessary during viral bioinseticides production scaling up, leads to the appearance of aberrant forms of virus, a process so called as "passage effect ". The main consequence of passage effect is the production of occlusion bodies (OB) decrease, preventing its production using in vitro process. In this study, it was carried out a serial passage of baculovirus Spodoptera frugiperda multiple nucleopolyhedrovirus, isolate 18, using Sf21 cells. A decrease in the production of occlusion bodies from 170 to 92 in the third to fourth passage was observed. A factorial experimental design (22) was employed to verify the influence of two input variables, concentration of the hormone 20 - hydroxyecdysone (CH) and cholesterol (CC) on the values of response variables (volumetric and the specific OB production) of the process, seeking to define the optimum operating ranges trying to reverse or minimize the passage effect. The result indicated a negative influence of the cholesterol addition and positive effect in the hormone supplementation which the optimum range found for the concentrations studied were 8 to 10μg/mL and 5 to 6.5 mg / mL, for cholesterol and hormone concentrations respectively. New experiments were performed with addition of hormone and cholesterol in order to check the influence of these additives on the OB production independently. While the best result obtained from the factorial experiment was 9.4 x 107 OB/mL and 128.4 specific OB/cell, with the addition of only 6μg/mL 20-hydroxyecdysone these concentrations increased to 1.9 x 108 OB/mL and 182.9 OB/cell for volumetric and specific OB production, respectively. This result confirms that the addition of the hormone 20-hydroxyecdysone enhances the SfMNPV in vitro production process performance using Sf21 cells
Dentre as pragas que atacam a cultura do milho no Brasil, destaca-se a Spodoptera frugiperda (J. E. Smith, 1797) (Lepidoptera: Noctuidae), conhecida no est?dio larval como lagarta-do-cartucho, considerada a praga chave da cultura, alimentando-se da planta em todas as suas fases de crescimento, principalmente dos cartuchos de plantas jovens. Para seu controle, tem-se empregado inseticidas qu?micos de amplo espectro, o que tem causado efeitos adversos ao homem e ao meio ambiente. Por essa raz?o, torna-se necess?ria a busca de alternativas mais eficientes, de baixo custo e de f?cil utiliza??o, como o uso de bioinseticidas, especialmente os baculov?rus. Devido ? instabilidade gen?tica durante a passagem seriada de baculov?rus em cultivo de c?lulas de inseto, o desenvolvimento da produ??o in vitro de bioinseticidas virais ? o maior desafio para a sua produ??o massal. Passagens sucessivas de v?rus usando v?rus extracelulares (BVs), necess?rias para o aumento de escala durante a produ??o de bioinseticida viral, leva ao aparecimento de formas aberrantes de v?rus, processo conhecido como efeito de passagem . A principal consequ?ncia do efeito passagem ? a diminui??o da produ??o de corpos de oclus?o (OB), inviabilizando economicamente sua produ??o pelo processo in vitro. Neste trabalho, foi realizada a passagem seriada do isolado 18 do baculov?rus Spodoptera frugiperda multiple nucleopolyhedrovirus em c?lulas Sf21. Foi observada uma queda na produ??o de OB de 170 para 92 da terceira para quarta passagem. Um planejamento experimental fatorial (22) foi empregado para verificar a influ?ncia de duas vari?veis de entrada, concentra??o de horm?nio 20- Hidroxiecdisona (CH) e concentra??o de colesterol (CC), sobre os valores das vari?veis de resposta do processo (produ??o volum?trica e produ??o espec?fica de OB), procurando definir as faixas ?timas de opera??o para reverter ou minimizar o efeito passagem. O resultado deste planejamento indicou influ?ncia negativa da adi??o do colesterol e positiva na adi??o de horm?nio, onde as faixas ?timas encontradas para as concentra??es estudadas foram: 8 a 10μg/mL e 5 a 6,5μg/mL, para as concentra??es de colesterol e horm?nio, respectivamente. Novos experimentos foram realizados com a adi??o individual de horm?nio e colesterol com a finalidade de verificar a influ?ncia destes adjuvantes na produ??o de OB de forma independente. No planejamento experimental obteve-se produ??o volum?trica de 9,4 x 107 OB/mL e espec?fica de 128,4 OB/c?lula. Quando se adicionou 6 μg/mL de 20-Hidroxiecdisona, as concentra??es elevaram-se para 1,9 x 108 OB/mL e 182,9 OB/c?lula, indicando que a adi??o do horm?nio melhorou a efici?ncia da produ??o in vitro de produ??o de SfMNPV em cultivos de c?lulas Sf21

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Funda??o de Amparo ? Pesquisa do estado de Minas Gerais (FAPEMIG)
Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior (Capes)
Lagartas da esp?cie Helicoverpa armigera foram identificadas no Brasil atacando extensas ?reas de produ??o agr?cola. Algumas caracter?sticas fazem deste inseto uma das principais pragas a sistemas agr?colas do mundo, o que torna este ataque um perigo eminente ?s ?reas de produ??o do pa?s. Estudos com os insetos coletados nestas ?reas s?o essenciais para que seja desenvolvido um manejo adequado. Estudos de laborat?rio podem selecionar poss?veis pr?ticas de manejo a fim de combater surtos de pragas no campo. Uma das formas de manejar o ambiente de forma segura ? adotar o Manejo Integrado de Pragas, onde se destaca o controle biol?gico utilizando baculov?rus. A lagarta de H. armigera ? suscept?vel a estirpes de baculov?rus, por?m ? necess?rio verificar esta suscetibilidade. Sendo assim, os objetivos deste trabalho foram selecionar estirpes aut?ctones de baculov?rus t?xicas as lagartas desta esp?cie. Foram realizados testes gen?ticos e biol?gicos entre as estirpes aut?ctones, comparando-as com o produto comercial Gemstar?, de origem norte americana. A an?lise comparativa do sequenciamento gen?tico realizada para os genes LEF-8 e LEF-9 revelaram que os isolados locais est?o, estreitamente, relacionados com esp?cies de baculov?rus da Austr?lia e da ?ndia. Todos os isolados testados possibilitaram o controle de lagartas de terceiro instar de H. armigera. Por?m, analises biol?gicas da concentra??o letal m?dia (CL50) e do tempo letal m?dio (TL50) variaram entre os isolados testados. O isolado HearNPV-BR2 apresentou os melhores resultados de CL50 e TL50. Ademais, este ? o primeiro registro da ocorr?ncia da esp?cie HearNPV-BR2 no Brasil, e suas propriedades inseticidas assinalam que a mesma pode ser ?til para a fabrica??o de bioinseticidas para o controle de H. armigera no pa?s.
Disserta??o (Mestrado) ? Programa de P?s-Gradua??o em Produ??o Vegetal, Universidade Federal dos Vales do Jequitinhonha e Mucuri, 2015.
ABSTRACT Larvea of Helicoverpa armigera species were identified in Brazil attacking extensive agricultural production areas. Some features make this one of the main insect pests in agricultural systems in the world, which makes this attack an imminent danger to agricultural production in the country. Studies with insects collected in these areas are essential for developed proper management. Laboratory studies can select possible management practices to combat pest outbreaks in the field. One way to manage the secure environment is to adopt the Integrated Pest Management, highlighting the biological control using baculovirus. The H. armigera larvea is susceptible to strains of baculovirus but it is necessary check this susceptibility. Therefore the objectives of this work were to select indigenous strains of toxic baculovirus larvea of this species. Genetic and biological tests were carried out between indigenous strains, comparing them with the commercial product Gemstar?, origin from North American (USA). Comparative analysis of genetic sequencing performed for the LEF-8 and LEF-9 genes, revealed that indigenous strains are closely related to species of baculovirus of the Australia and India. All strains tested allowed control of the third instar of the H. armigera larvea. However, biological analysis of median lethal concentration (LC 50) and median lethal time (LT50) varied among the strains tested. The HearNPV-BR2 strain showed the best results of CL50 and TL50. Moreover, this is the first record of the occurrence of HearNPV-BR2 species in Brazil and its insecticidal properties indicate that BR2 may be useful for the manufacture of biological insecticides for H. armigera control in the country.

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Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico
In vivo production of viral biopesticides is the major source of viral insecticides currently in the marketplace. However, this system presents limitations during production scale-up. For the Spodoptera frugiperda nucleopolyhedrovirus (SfMNPV), the insect used for replication has cannibalistic characteristics, thus production is even more difficult. Insect cells are commonly used for in vitro baculovirus production. Most of these cell lines are derived from Lepidoptera species. The Sf21 cell line is derived from Spodoptera frugiperda caterpillar ovarian tissue, and its clonal isolate Sf9 has been used for biopesticide production due to its ease of growth in suspension cultures. In this work, the in vitro production capabilities of a Brazilian SfMNPV isolate obtained from cornfields was evaluated. Comparison of polyhedra production was carried out using both Sf21 and Sf9 cells, based on volumetric and specific yields. Both cell lines were cultivated in Hyclone medium supplemented with different fetal bovine serum concentrations (2,5 and 5%). The best results were obtained using Sf9 cells supplemented with 5% serum. These results were further confirmed quantitively through kinetic parameter estimation for both cells lines and different serum concentrations. After seven successive passages, this system still presented high specific polyhedra production
A produ??o in vivo de biopesticidas virais ? a maior fonte destes inseticidas presentes no mercado atualmente. Entretanto, o sistema in vivo apresenta limita??o em rela??o ao escalonamento de produ??o. No caso do v?rus Spodoptera frugiperda nucleopoliedrovirus (SfMNPV), o inseto usado para sua multiplica??o tem caracter?sticas canibais, o que dificulta ainda mais a sua produ??o in vivo. As c?lulas de inseto s?o comumente utilizadas para produ??o in vitro de baculov?rus. V?rias linhagens destas c?lulas s?o derivadas principalmente da esp?cie Lepidoptera. A linhagem Sf21 ? derivada do tecido ovariano da lagarta Spodoptera frugiperda e um clone isolado da linhagem original, denominado Sf9, tem sido utilizado para produ??o de biopesticidas, por apresentar f?cil crescimento em cultivo em suspens?o. Neste trabalho, foi testada a viabilidade de produ??o in vitro de um isolado viral brasileiro de SfMNPV obtido em lavouras de milho. A produ??o de poliedros, em c?lulas Sf21 e Sf9, foi determinada com base na avalia??o comparativa da produtividade volum?trica e espec?fica destes poliedros. Ambas as linhagens celulares foram cultivadas, em suspens?o, em meio HyClone suplementado com diferentes concentra??es de soro fetal bovino (2,5 e 5%). A c?lula Sf9 suplementada com 5% de soro apresentou os melhores resultados de produ??o. Os resultados foram confirmados quantitativamente atrav?s dos par?metros cin?ticos estimados para as duas linhagens e diferentes concentra??es de soro. O sistema demonstrou, ap?s sete passagens sucessivas, uma alta produ??o espec?fica de poliedros

Global large-scale adoption of Bt transgenic crops has provided effective management of key insect pests and have greatly reduced insecticide use. However, some field populations of several insect species have evolved resistance to Bt crops in the field, which threatens the continuing success of Bt crops. The cotton bollworm (Helicoverpa zea) is among the first pest reported to have field-evolved resistance to Cry1Ac and Cry2Ab, but the underlying mechanisms remain largely unknown. To determine the current resistance status of the field populations of H. zea and to elucidate the mechanisms of Bt resistance in this pest, I conducted a series of experiments including bioassays of field populations as well as biochemical and molecular comparisons of midgut proteases and putative Cry1Ac receptors between Cry1Ac-susceptible and -resistant strains. Diet incorporation bioassays of six field populations of H. zea collected from Tifton, Georgia USA in 2008 and 2009 indicated that, comparing to LAB-S, a susceptible laboratory strain, all six field populations were significantly resistant to Cry1Ac toxin and one of three field strains was significantly resistant to Cry2Ab toxin. Across the five populations, survival on leaf-discs producing Cry1Ac was positively correlated to the lethal concentration that kills 50% of the population (LC₅₀) for Cry1Ac from diet bioassays. These results support previous findings of field-evolved resistance to Bt crops in H. zea and suggest an overall increase in resistance to Cry1Ac from 2002 to 2009.One of the six field population, which was designed as GA and had 55-fold resistance to Cry1Ac, was further selected with Cry1Ac in the laboratory to generate a more resistant strain, which was designated as GA-R and had 560-fold resistance to Cry1Ac. Total protease activity of the midgut extracts from GA-R and GA strains is significantly lower than that from the susceptible laboratory strain LAB-S. Among the proteases contributing to the total activity, trypsin-like and chymotrypsin-like activities of GA-R and GA midgut extracts are significantly lower than that from the susceptible strain, while no difference in elastase-like activity is evident. Decreased proteolytic activity was correlated to the decreased Cry1Ac activation rate of midgut extracts of the GA-R and GA strains. Cytotoxicity assays with H. zea midgut cells show that the product of Cry1Ac protoxin digested with GA-R and GA midgut extracts has significantly lower cytotoxicity when compared with that digested with the susceptible strain midgut extracts. Transcriptional analysis of a limited number of protease genes did not identify specific proteases involved in the decline in Cry1Ac activation in GA-R and GA. These results indicate that the decreased Cry1Ac activation rate by midgut proteases is involved in the field originated Cry1Ac resistance in the H. zea GA-R and GA strains. I also compared the cDNA sequences and expression levels of the putative Cry1Ac receptors cadherin, aminopeptidase 1 (APN1), alkaline phosphatase 2 (ALP2) and ATP-binding cassette subfamily C member 2 (ABCC2) in LAB-S and GA-R. No indels (insertions and deletions) were found in the cDNA sequences of the resistant alleles of the four receptors, relative to those of the susceptible alleles. While there were no amino acid point mutations in the resistant alleles of ALP2 and ABCC2, we found 2 and 14 consistent amino acid point mutations in the resistant alleles of cadherin and APN1, respectively. However, neither cadherin nor APN1 point mutations were genetically linked to Cry1Ac resistance in GA-R. Quantitative RT-PCR analysis revealed no differences in the transcripts of the four receptors between the two strains. Taken together, these results indicate that the four receptors are not involved in Cry1Ac resistance in the GA-R strain of H. zea.

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No gênero Thrichomys cinco descrições de espécies são reconhecidas, sendo que geralmente este gênero é considerado monotípico. Thrichomys apereoides (Lund, 1839) descrito de Lagoa Santa em Minas Gerais é um roedor echimídeo que difere da maioria dos roedores desta família por apresentar pelagem macia e cauda densamente pilosa. Em estudos recentes que enfocam a espécie, os resultados das análises já realizadas (morfométricas, citogenéticas e moleculares) sugerem que Thrichomys seja politípico. O objetivo deste estudo foi realizar análises morfológicas (crânio e báculo) e análise citogenética em três amostras populacionais de Thrichomys apereoides em Minas Gerais (Lagoa Santa, Matozinhos e Santana do Riacho) incluindo os espécimes coletados por Lund (1839), na Bahia (Morro do Chapéu, Mucugê, Palmeiras e Rio de Contas) e em Mato Grosso (Barão de Melgaço), visando compreender os limites morfológicos de possíveis unidades evolutivas independentes dentro do gênero Thrichomys. As análises qualitativa e quantitativa do crânio foram úteis na diferenciação das amostras. Estados de caracteres cranianos, Análise das Funções Discriminantes e Análise dos Componentes Principais discriminaram as amostras. A amostra da Bahia foi totalmente discriminada no eixo canônico principal 1 e parcialmente discriminada das demais amostras no componente principal 1. A análise bacular não discriminou as amostras. A análise citogenética apresentou cariótipos diferenciados: Minas Gerais com 2n = 28 e NF = 50; Bahia com 2n = 26 e NF = 48; Mato Grosso com 2n = 34 e NF = 64. A amostra de Minas Gerais é alocada a Thrichomys aperoides. A amostra da Bahia é alocada a Thrichomys inermis. A amostra de Mato Grosso é alocada a Thrichomys pachyurus.
Five nominal forms in Thrichomys has described, but this genera is currently regarded as monotypic. Thrichomys apereoides (Lund, 1839) described from Lagoa Santa (Minas Gerais) can be distinguished from other genera within the Echimyidae by its soft fur and densely-haired tail. Morphometric, karyological and molecular analysis in Thrichomys suggested that genera is politypic. The present study realized morphological (cranial and bacular) and Karyological analysis in three Thrichomys apereoides samples from Minas Gerais (Lagoa Santa, Matozinhos and Santana do Riacho) within specimens Lund ( 1839) collected, Bahia (Morro do Chapéu, Mucugê Palmeiras and Rio de Contas) and Mato Grosso (Barão de Melgaço) attempt to found independent evolutionary units. Qualitative and quantitative cranial analysis shows differences among the samples studied. Cranial traits conditions, Discriminant Functions Analysis and Principal Component Analysis discriminated the samples. The sample from Bahia has fully discriminated a1ong an axis CV 1 and parti11y discriminated along axis CP 1 . Bacular analysis not discriminated the samples. Karyological analysis discriminated the samples: 2n = 28 and FN = 50 from Minas Gerais; 2n = 26 and FN = 48 from Bahia; 2n = 34 and NF = 64 from Mato Grosso. Thrichomys apereoides is the oldest name the sample from Minas Gerais. Thrichomys inermis is the oldest name the sample from Bahia. Thrichomys pachyurus is the oldest name the sample from Mato Grosso.

[For the Abstract, please see the PDF files below, namely "front.pdf"] CONTENTS. Chapter 1 Introduction. Chapter 2 Gross and microscopic visceral anatomy of the male Cape fur seal with reference to organ size and growth. Chapter 3 Age determination and growth in the male Cape fur seal: part one, external body. Chapter 4 Age determination and growth in the male Cape fur seal: part two, skull. Chapter 5 Age determination and growth in the male Cape fur seal: part three, baculum. Chapter 6 Suture age as an indicator of physiological age in the male Cape fur seal. Chapter 7 Sexual dimorphism in the adult Cape fur seal: standard body length and skull morphology. Chapter 8 Reproduction in the male Cape fur seal: age at puberty and annual cycle of the testis. Chapter 9 Diet and foraging behaviour of the Cape fur seal. Chapter 10(a) The Impact of the fur seal industry on the distribution and abundance of Cape fur seals. Chapter 10(b) South African Airforce wildlife rescue: Cape fur seal pups washed from Black Rocks, Algoa Bay, during heavy seas, December 1976. Chapter 11(a) Operational interactions between Cape fur seals and fisheries: part one, trawl fishing. Chapter 11(b) Operational interactions between Cape fur seals and fisheries: part two, squid jigging and line fishing. Chapter 11(c) Operational interactions between Cape fur seals and fisheries: part three, entanglement in man-made debris. Chapter 12 Concentrations of heavy metals (Cd, Cu, Pb, Ni & Zn) and organochlorine contaminants (PCBs, DDT, DDE & DDD) in the blubber of Cape fur seals. Chapter 13 Endoparasites of the Cape fur seal. Chapter 14(a) Preliminary investigations of shark predation on Cape fur seals. Chapter 14(b) Aggressive behaviour of an adult male Cape fur seal towards a great white shark Carcharodon carcharias. Chapter 15 Conclusions and future directions.

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Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior (CAPES)
O uso de agrot?xicos em cultivo agr?cola ? cada vez maior. Os inseticidas qu?micos usados na agroind?stria causam grandes preocupa??es como a contamina??o ambiental e a falta de seletividade aos organismos. Uma alternativa para esse problema ? a utiliza??o de bioinseticidas. Os bioinseticidas s?o mais seguros que os agrot?xicos comuns e apresentam vantagens como menor toxicidade e alta especificidade. Ademais, podem ser utilizados em menor quantidade, decomp?em-se mais r?pido e sua utiliza??o associada com os inseticidas sint?ticos faz com que o uso de agrot?xicos seja menor, causando baixo impacto ambiental. Em particular, bioinseticidas do tipo baculov?rus t?m sido apontados como uma op??o de substitui??o a estes inseticidas qu?micos na agricultura. Neste trabalho, foi realizada a produ??o in vitro do baculov?rus Spodoptera frugiperda nucleopolyhedrovirus e o estudo do seu perfil eletrofor?tico com base nas prote?nas virais presentes nos v?rus extracelulares deste baculov?rus. As part?culas de v?rus extracelulares obtidas durante a passagem seriada do baculov?rus foram analisadas por eletroforese em gel desnaturante e, como esperado, a poliedrina - prote?na respons?vel pela forma??o do corpo de oclus?o - com aproximadamente 30 kDa foi destaque em todas as passagens analisadas. Prote?nas na regi?o entre 14,4 e 97 kDa tamb?m foram detectadas. A an?lise do efeito da adi??o do horm?nio ?-ecdisona extra?do do Ginseng brasileiro e do horm?nio ecdisona sint?tico na produ??o do baculov?rus Spodoptera foi realizada na terceira e quarta passagens em c?lulas Sf21, momento em que a forma??o dos corpos de oclus?o foi reduzida. Os resultados mostraram que o horm?nio ecdisona n?o influenciou na produ??o dos corpos de oclus?o na terceira passagem. Por?m, na quarta passagem ao se adicionar o horm?nio ecdisona, tanto o sint?tico como o extra?do do Ginseng brasileiro, houve um aumento aproximado de duas vezes na forma??o dos corpos de oclus?o quando comparados ? produ??o sem adi??o de ecdisona. Utilizando este processo de infec??o, foi realizada uma modelagem e simula??o matem?tica que representasse o mecanismo de produ??o do bioinseticida.
O uso de agrot?xicos em cultivo agr?cola ? cada vez maior. Os inseticidas qu?micos usados na agroind?stria causam grandes preocupa??es como a contamina??o ambiental e a falta de seletividade aos organismos. Uma alternativa para esse problema ? a utiliza??o de bioinseticidas. Os bioinseticidas s?o mais seguros que os agrot?xicos comuns e apresentam vantagens como menor toxicidade e alta especificidade. Ademais, podem ser utilizados em menor quantidade, decomp?em-se mais r?pido e sua utiliza??o associada com os inseticidas sint?ticos faz com que o uso de agrot?xicos seja menor, causando baixo impacto ambiental. Em particular, bioinseticidas do tipo baculov?rus t?m sido apontados como uma op??o de substitui??o a estes inseticidas qu?micos na agricultura. Neste trabalho, foi realizada a produ??o in vitro do baculov?rus Spodoptera frugiperda nucleopolyhedrovirus e o estudo do seu perfil eletrofor?tico com base nas prote?nas virais presentes nos v?rus extracelulares deste baculov?rus. As part?culas de v?rus extracelulares obtidas durante a passagem seriada do baculov?rus foram analisadas por eletroforese em gel desnaturante e, como esperado, a poliedrina - prote?na respons?vel pela forma??o do corpo de oclus?o - com aproximadamente 30 kDa foi destaque em todas as passagens analisadas. Prote?nas na regi?o entre 14,4 e 97 kDa tamb?m foram detectadas. A an?lise do efeito da adi??o do horm?nio ?-ecdisona extra?do do Ginseng brasileiro e do horm?nio ecdisona sint?tico na produ??o do baculov?rus Spodoptera foi realizada na terceira e quarta passagens em c?lulas Sf21, momento em que a forma??o dos corpos de oclus?o foi reduzida. Os resultados mostraram que o horm?nio ecdisona n?o influenciou na produ??o dos corpos de oclus?o na terceira passagem. Por?m, na quarta passagem ao se adicionar o horm?nio ecdisona, tanto o sint?tico como o extra?do do Ginseng brasileiro, houve um aumento aproximado de duas vezes na forma??o dos corpos de oclus?o quando comparados ? produ??o sem adi??o de ecdisona. Utilizando este processo de infec??o, foi realizada uma modelagem e simula??o matem?tica que representasse o mecanismo de produ??o do bioinseticida.

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Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES
The vitamin D receptor (VDR) is a cytoplasmic transcription factor and when activated by its ligand, translocates to the nucleus and interacts with DNA in the promoter regions with which has affinity, acting as a modulator of gene transcription and thereby producing multiple biological effects. Its main activities are the regulation and maintenance of plasma levels of calcium and phosphorus, as well as presenting immunomodulatory activities, such as the suppression of T cell activation, formation of secretion patterns of cytokines, modulation of proliferation and interference in the apoptosis. So this receptor is an important target for drugs that can help in the discovery of new immunomodulators. The present work had the objective to produce the recombinant vitamin D receptor in its soluble form for conducting future assays of drug screening of potential immunomodulators and drug-receptor interaction studies. For this, we used initially Escherichia coli expression system transformed with the plasmid HS_VDR_EC1-PQE T7, but it was only possible to obtain the protein in the insoluble fraction, even varying the temperature, time of induction and IPTG concentration. In an attempt to obtain soluble VDR, we used a eukaryotic expression system in insect cells using the baculovirus as a vector. It was built a vector, pFASTBacHT_VDR, which had the sequence of this protein cloned from pCMX.VDR. From there, it was possible to obtain recombinant bacmids used in transfection of insect cells, generating recombinant baculovirus, to then proceed with the expression of VDR.
O receptor de vitamina D (VDR) ? um fator de transcri??o g?nica citoplasm?tico e, quando ativado pelo seu ligante, transloca-se para o n?cleo e interage com regi?es promotoras no DNA com a qual apresenta afinidade, atuando como fator modulador da transcri??o g?nica e assim produzindo m?ltiplos efeitos biol?gicos. Suas principais atividades s?o a regula??o e a manuten??o dos n?veis plasm?ticos de c?lcio e f?sforo, e apresenta atividades imunomoduladoras, como a supress?o da ativa??o de c?lulas T, forma??o de padr?es de secre??o de citocinas, a modula??o da prolifera??o e interfer?ncia na apoptose. Sendo assim, esse receptor representa um importante alvo de drogas que pode contribuir na descoberta de novos f?rmacos com a??o imunomoduladora. O presente trabalho teve, como objetivo, produzir o VDR recombinante na forma sol?vel para a realiza??o de futuros ensaios de triagem de potenciais drogas com a??o imunomoduladora e estudos de intera??o droga-receptor. Para isso, foi utilizado, inicialmente, o sistema de express?o em Escherichia coli, utilizando o plasm?deo HS_VDR_EC1-PQE T7, por?m s? foi poss?vel obter a prote?na na fra??o insol?vel, mesmo variando a temperatura, o tempo de indu??o e a concentra??o de IPTG. Na tentativa de obter o VDR sol?vel, foi utilizado um sistema de express?o eucari?tico em c?lulas de inseto, utilizando como vetor o baculov?rus. Foi constru?do um vetor pFASTBacHT_VDR, o qual teve a sequ?ncia desta prote?na clonada a partir do pCMX.VDR. A partir da?, foi poss?vel obter bacm?deos recombinantes, utilizados na transfec??o de c?lulas de inseto, gerando baculov?rus recombinantes, para, ent?o, seguir com a express?o do VDR.

L’objectif de ce projet de thèse est d’étudier et caractériser les mécanismes impliqués dans la bascule respiro-fermentaire chez des cellules eucaryotes dotées d’un métabolisme mitochondrial. Les cellules eucaryotes ont des besoins différents en oxygène pour la production d’énergie et leur survie dans un environnement donnée. Elles sont qualifiées de type aérobie stricte lorsque la présence d’oxygène leur est nécessaire ou aéro-anaérobie facultatif dans le cas où l’oxygène n’est pas indispensable à la production d’énergie. La levure Yarrowia lipolytica a été choisie comme modèle d’étude de par sa particularité à être un micro-organisme aérobie strict avec une grande capacité d’accumuler de lipides et de production d’acides organiques. Les études expérimentales et analytiques, par l’emploi de méthodes mathématiques de modélisation du métabolisme, ont permis d’identifier des contraintes métaboliques impliquées dans la transition respiro-fermentaire chez cette levure au métabolisme énergétique oxydatif. La production de l’acide citrique par Y. lipolytica, déjà rapportée dans la littérature, a été choisi comme un marqueur de cette transition respiro-fermentaire. Nous avons découvert que l’inhibition de la protéine oxydase alternative (AOX), impliquée dans la respiration mitochondriale, par la molécule n-propyl gallate (nPG) permet d’améliorer le rendement de production d’acide citrique par fermentation du glucose dans une culture de Y. lipolytica. Ces résultats montrent que la nPG, déjà utilisée dans l’industrie agro-alimentaire et pharmaceutique en tant que conservateur joue sur la bascule respiro-fermentaire par inhibition de la consommation d’oxygène et stimule ainsi la production d’acide citrique. La modélisation du réseau métabolique de Y. lipolytica, décrit à l’échelle du genome, par dynamic Flux Balance Analysis (dFBA) a permis d’identifier l’accumulation des espèces oxydantes dites ROS (Reactive Oxygen Species) comme un levier majeur de la bascule respiro-fermentaire et donc de la production d’acide citrique chez la levure Y. lipolytica. De plus, nos résultats préliminaires montrent que l’oxydation des lipides accumulés par Y. lipolytica pourrait être à l’origine de la génération des ROS. Cette étude doit être approfondie expérimentalement et constitue un apport important pour l’industrie agro-alimentaire et pharmaceutique.Mots clés : Bascule respiro-fermentaire, Acide citrique, lipides, Yarrowia lipolytica, n-propyl gallate, Reactive Oxygen Species, modélisation, dynamic Flux Balance Analysis
The main goal of this thesis project is to study and characterize mechanisms involved in respiratory to fermentative shift in eukaryotic cells endowed with mitochondrial metabolism. Eukaryotic cells have different oxygen requirements for energy production and survival in a given environment. They are described as strict aerobic when the presence of oxygen is necessary or optional aero-anaerobic in when oxygen is not essential for energy production. The yeast Yarrowia lipolytica was chosen as our study model thanks to its particularity since it is a strict aerobic microorganism with a high capacity to accumulate lipids and to produce organic acids. Experimental and analytical studies, using mathematical methods for modeling cell metabolism, allowed us to identify metabolic constraints involved in respiratory to fermentative transition in this yeast showing oxidative energy metabolism. Production of citric acid by Y. lipolytica, already reported in the literature, has been chosen as a marker for this in respiratory to fermentative shift. We found that the inhibition of the alternative oxidase protein (AOX) involved in mitochondrial respiration, by adding n-Propyl gallate (nPG) molecule improves the yield of citric acid production by fermentation of glucose in a Y. lipolytica culture. These results show that nPG, already used in food and pharmaceutical industry as a preservative, plays on respiratory to fermentative balance by inhibition of oxygen consumption and thus stimulates the production of citric acid. Modeling of the metabolic network of Y. lipolytica, described at genome-scale, by dynamic Flux Balance Analysis (FBA) has identified the accumulation of intracellular ROS (Reactive Oxygen Species) species as major levers for respiratory to fermentative shift and therefore the production of citric acid by Y. lipolytica. Therefore, our preliminary results show that oxidation of lipids accumulated by Y. lipolytica could be involved in generation of ROS species. This study must be experimentally deepened and constitutes an important contribution for the agri-food and pharmaceutical industry.Key words: Respiratory to fermentative shift, Citric acid, lipids, Yarrowia lipolytica, n-Propyl gallate, Reactive Oxygen Species, modeling, dynamic Flux Balance Analysis

This dissertation examines the use of argumentum ad baculum in preaching in general and the sermons of George Whitefield in particular. Argumentum ad baculum has traditionally been considered an informal fallacy of relevance. The fallacy can be defined as an appeal to force or an appeal to fear. Chapter 1 discusses the relationship of argumentum ad baculum with the empirical study of fear appeals and the rhetorical use of pathos. Attention is also given to the preaching of Whitefield and his place in the history of preaching as an innovator. Whitefield's role in the shift to a more passionate and emotional sermon style is noted. The chapter also addresses the challenges a study of Whitefield's sermons presents. Chapter 2 is devoted to defining argumentum ad baculum, examining the history of the phrase, the two ways it has been defined, the nature of it as a fallacy, and fear appeals as a part of the definition. The chapter includes a discussion of source credibility in relation to fear appeals. Chapter 3 analyses the sermons of Whitefield to identify his use of fear appeals. The types of fear appeals he used in his sermons are listed along with evidentiary sermon material. The types of material Whitefield used to formulate the appeals are also discussed. Chapter 4 gives attention to the effect of Whitefield's fear appeals on his auditors. In order for an appeal to be effective, it must first arouse fear in the recipients of the appeal. Historical narratives are examined from Whitefield himself, eyewitness accounts, and personal testimonies of those who were present at his meetings. The chapter provides evidence of the general and specific effect of Whitefield's fear appeals. Chapter 5 concerns the ethicality of Whitefield's appeals. The chapter surveys a number of standards for ethical judgment. The chapter argues that Whitefield's use of fear in his published sermons was ethical, primarily because of the intention with which he used them. Chapter 6 offers guidelines for the contemporary use of argumentum ad baculum in preaching. Modern audiences are unaccustomed to the use of fear for persuasive means. However, this type of argumentation can be used ethically and effectively.

This study is focused on a bacular development in Microtus arvalis, histological sections in two species of voles for the detection of a presence of os clitoridis, and description of an os clitoridis in Sciurus vulgaris. in the last part, there is the penile morphology and bacular structure of one species of African mole rat (Heliophobius argenteocinereus) described.

This master thesis consists of two main parts. The first of them represents a compilation of information and facts about baculum, specifically about its presence, function and evolution in mammals. Further I focused on biological characters of five genera of African muroid rodents (Acomys, Aethomys, Gerbilliscus, Saccostomus, Stenocephalemys), which were analyzed in detail in the second part of my thesis in respect of their penial and bacular morphology.

Tese de mestrado, Bioquímica, Universidade de Lisboa, Faculdade de Ciências, 2009
As hidrogenases são enzimas que catalisam a oxidação reversível de H2, considerado um combustível alternativo limpo. Como a sua inibição por O2 tem sido um dos grandes obstáculos à sua aplicação em sistema biotecnológicos, as poucas hidrogenases resistentes ou tolerantes ao O2 têm despertado muita curiosidade. O centro activo das hidrogenases do tipo [NiFe] contém um ferro e um níquel. Este último é coordenado por quatro cisteínas. O subgrupo das hidrogenase [NiFeSe], onde uma das cisteínas está substituída por uma selenocisteína, é particularmente interessante porque apresenta tolerância ao O2 e maior eficiência na produção de H2 quando comparada com as hidrogenases [NiFe] padrão, não se sabe, no entanto, como é que o selénio influencia estas propriedades. A cristalografia de raios X revelou a presença de canais que permitem ao H2 e O2 aceder ao centro activo, conservados nas hidrogenases padrão. No caso das hidrogenases reguladoras existem indícios de que o estreitamento destes canais, e consequente redução do acesso dos gases, pode ser a razão para a sua tolerância ao O2. Na hidrogenase [NiFeSe] de Dm. baculatum um dos canais identificado nas hidrogenases padrão não está conservado. O estudo apresentado neste trabalho tem por objectivo analisar os processos de difusão na hidrogenase [NiFeSe] de Dm. baculatum, através de simulações de dinâmica molecular e compará-los com o reportado para hidrogenase [NiFe] de D. gigas, de modo a perceber se estes podem influenciar a diferença de desempenho destes dois enzimas. Os resultados indicam que a quantidade de H2 interiorizada, assim com a sua densidade junto ao níquel é superior na hidrogenase [NiFeSe] de Dm. baculatum. As três principais vias de acesso são similares nas duas hidrogenases, mas a hidrogenase [NiFeSe] apresenta maior afluência no canal da subunidade pequena e menor circulação de H2 na zona do canal menos conservado.
Hydrogenases catalyze the reversible oxidation of H2 to protons and electrons. These enzymes have large potential for producing and oxidizing H2, which is considered an alternative green fuel. The O2 inhibition is an obstacle to their application in biotechnological systems. Nonetheless, there are some hydrogenases that are O2 resistant or tolerant. The [NiFe]-type hydrogenases have a bimetallic active site with an iron and a nickel ion. In the subgroup of [NiFeSe]-hydrogenases, one of the four cysteins that coordinate the nickel in [NiFe]-hydrogenases, is replaced by a selenocystein. These [NiFeSe]- hydrogenases are particularly interesting for their higher H2 production rate when compared to the standard [NiFe]-hydrogenases, in addition to their O2 tolerance. It is not clear which role selenium plays in these properties. In regulatory hydrogenases, the diminished gas diffusion has been pointed by some studies as the reason for their O2 resistance, suggesting that the hydrophobic channels that provide gas access in hydrogenases are narrower in these enzymes. One of those channels identified in the Xray structure of standard [NiFe]-hydrogenase appears to be less conserved in the NiFeSe]-hydrogenase of Desulfomicrobium baculatum. The influence of H2 diffusion in the Dm. baculatum [NiFeSe]-hydrogenase particular properties is addressed in this work. The H2 diffusion in this protein is simulated with molecular dynamics methods, and the results are compared with a previous study concerning the Desulfovibrio gigas [NiFe]-hydrogenase. The results indicate that the hydrogen internalization is higher for the [NiFeSe]- hydrogenase and a higher H2 density near the nickel ion is registered for these enzymes. Although the three main pathways are roughly maintained, the flow of H2 in the small subunit channel appears to be augmented in the [NiFeSe]-hydrogenase, while the least conserved channel in the X-ray structure present a reduced flow.

Chapter 1 Introduction. Chapter 2 Gross and microscopic visceral anatomy of the male Cape fur seal with reference to organ size and growth. Chapter 3 Age determination and growth in the male Cape fur seal: part one, external body. Chapter 4 Age determination and growth in the male Cape fur seal: part two, skull. Chapter 5 Age determination and growth in the male Cape fur seal: part three, baculum. Chapter 6 Suture age as an indicator of physiological age in the male Cape fur seal. Chapter 7 Sexual dimorphism in the adult Cape fur seal: standard body length and skull morphology. Chapter 8 Reproduction in the male Cape fur seal: age at puberty and annual cycle of the testis. Chapter 9 Diet and foraging behaviour of the Cape fur seal. Chapter 10(a) The Impact of the fur seal industry on the distribution and abundance of Cape fur seals. Chapter 10(b) South African Airforce wildlife rescue: Cape fur seal pups washed from Black Rocks, Algoa Bay, during heavy seas, December 1976. Chapter 11(a) Operational interactions between Cape fur seals and fisheries: part one, trawl fishing. Chapter 11(b) Operational interactions between Cape fur seals and fisheries: part two, squid jigging and line fishing. Chapter 11(c) Operational interactions between Cape fur seals and fisheries: part three, entanglement in man-made debris. Chapter 12 Concentrations of heavy metals (Cd, Cu, Pb, Ni & Zn) and organochlorine contaminants (PCBs, DDT, DDE & DDD) in the blubber of Cape fur seals. Chapter 13 Endoparasites of the Cape fur seal. Chapter 14(a) Preliminary investigations of shark predation on Cape fur seals. Chapter 14(b) Aggressive behaviour of an adult male Cape fur seal towards a great white shark Carcharodon carcharias. Chapter 15 Conclusions and future directions.