Lysine methylation is a post-translational modification that regulates a large array of functionally diverse processes that are vital for cellular function. The role of methylation is best characterized on histone proteins due to their high concentration in the cell, but alongside histone modifications, lower abundance non-histone methylation is emerging as a prevalent and functionally diverse regulator of cellular processes. The direct biological impact of non-histone lysine methylation is less well understood because they are difficult to detect. The dynamic concentration range of the proteome masks their signal during proteomic analysis which impedes the detection of these low abundance methylated proteins. Increasing the concentration of proteins bearing methylation is required for improved discovery. This requires enriching the post-translational modification with a capturing reagent prior to analysis. This thesis details an optimized method for using the supramolecular host p-sulfonatocalix[4]arene as a stationary phase methyllysine enrichment reagent for real-life cell-extracted proteins. Prior to the optimizations described in this thesis, cell-derived peptide extracts were not retained within an early generation upper-rim modified calixarene column. But with the new protocols detailed in this thesis, proteins extracted from both cultured prostate cancer cells and industrially sourced brewer’s yeast were successfully retained by a lower-rim modified calixarene column. Thousands of methylated proteins with diverse functions and cellular localization were discovered using this method. Detection of low abundance methylated proteins will aid our discovery of all cellular methylation marks, which in turn, will help delineate their biological functions. / Graduate / 2020-11-30

Submitted by Maria Medeiros (maria.dilva1@ufcg.edu.br) on 2018-11-06T11:40:02Z No. of bitstreams: 1 LUCIANO FERNANDES MONTEIRO - TESE (PPGEP) 2012.pdf: 1345638 bytes, checksum: 56948a137db5ba5ecb07f475b625a828 (MD5) / Made available in DSpace on 2018-11-06T11:40:02Z (GMT). No. of bitstreams: 1 LUCIANO FERNANDES MONTEIRO - TESE (PPGEP) 2012.pdf: 1345638 bytes, checksum: 56948a137db5ba5ecb07f475b625a828 (MD5) Previous issue date: 2012-05-11 / Capes / As indústrias de processamento de frutas geram grandes quantidades de resíduos e em geral estes resíduos são descartados no ambiente, misturando os oriundos das diferentes frutas. Um aproveitamento racional e eficiente desses resíduos como substrato para a produção de proteínas microbianas poderá dar resultados satisfatórios na produção de suplementos para rações, contribuindo também para minimizar os problemas de deposição dos resíduos e agregar valor aos resíduos em forma de suplemento proteico. O presente trabalho foi realizado com o objetivo geral de viabilizar o processo de obtenção de enriquecidos proteicos para ração animal a partir de mistura dos resíduos agroindustriais do processamento de frutas, com estudos de casos com os resíduos do abacaxi, pedúnculo do caju e maracujá visando a produção de suplemento proteico animal, a logística de distribuição e a minimização dos custos do produto final através de análise econômica. Foi realizada a caracterização físicoquímica da mistura dos resíduos in natura e enriquecidos. O estudo do enriquecimento proteico foi realizado por fermentação semisólida (FSS) utilizando a levedura Saccharomyces cerevisiae como inóculo e mistura de resíduos de abacaxi, caju e maracujá como substrato. O enriquecimento foi realizado em estufa com circulação de ar e em câmara climática com umidade relativa do ar controlada. As condições de operação foram: percentagem de leveduras entre 3 e 8%, temperatura de 33 e 35 ºC e umidade relativa do ar de 60, 70 e 80%. Foi observado que a mistura de resíduos, mesmo em condições diversas das recomendadas, apresentou índice de enriquecimento proteico compatível com os relatados na literatura para os resíduos individuais, obtendo-se um produto com 20% de proteína bruta (b.s.). As melhores condições foram T = 35 ºC, inóculo de 3% de levedura e 80% de umidade relativa. Nestas condições o consumo de açúcares redutores foi de 3,422% e a umidade final dos resíduos enriquecidos de 78,24%. Foi realizado levantamento para identificação dos fornecedores de resíduos e de seus potenciais produtores na Região Nordeste, especificamente nos estados de Alagoas-AL, Ceará-CE, Paraíba-PB, Pernambuco-PE, Rio Grande do Norte-RN e Sergipe-SE. Mediante estudo de caso, através do método do centro de gravidade a localização recomendada para uma unidade produtora no estado de Sergipe foi o município de Estância. A análise preliminar de custos aponta para a viabilidade de produção, com preço final aproximado de R$ 0,47 por quilograma de suplemento proteico produzido, o que indica um valor muito competitivo em relação às alternativas tradicionais. / The fruit processing industries generate large quantities of wastes and the mixing of residuals from different fruits are usually disposed in the environment. A rational and efficient use of these wastes as a substrate for microbial protein enrichment may give satisfactory results in the production of supplements to diets obtaining products with value added. This use of residuals also contributes to minimize waste disposal problems. This work was carried out to optimize the overall process of obtaining enriched protein supplements for animal feed using the mixture of agro industrial wastes from processed fruits. Case studies with pineapple shell, cashew-apple bagasse and passion fruit shell and albedo aimed a better storage, distribution logistics and minimizing the costs of the final product through economic analysis. Fresh and enriched residuals mixture was characterized by their physicochemical properties. The study of protein enrichment was performed by solid state fermentation (SSF) using the yeast Saccharomyces cerevisiae as inoculum and a 1:1:1 mixture of pineapple shells, passion fruit shells and albedo and cashew apple bagasse as substrate. The enrichment was carried out in an oven with air circulation and in a climatic chamber with controlled relative humidity. The operating conditions were: yeast percentage between 3 and 8%, temperature 33ºC and 35 ° C and relative humidity between 60 and 80%. It was found that the mixture of wastes, even when processed out of the optimal conditions recommended for each one, presented protein enrichment index compatible with those reported in literature for the individual waste , obtaining a product with 20% crude protein (b.s). In the studied range it was found that the best conditions were T = 35 ° C, 3% of yeast inoculum and 80% relative humidity. Under these conditions the consumption of reducing sugars was 3.422% and the final moisture content of enriched waste 78.24%. Survey was conducted to identify the suppliers of waste and its potential producers in the Northeast of Brazil, specifically at the States: AL, CE, PB, PE, RN and SE. Through case study, by the method of center of gravity it was found that the neighbors of Estancia is the recommended location for a production unit in the State of Sergipe. A preliminary cost analysis indicates the feasibility of production, with the final price of approximately R$ 0.47 per kilogram of protein supplement produced, which indicates a very competitive value compared to traditional alternatives.

Submitted by Maria Medeiros (maria.dilva1@ufcg.edu.br) on 2018-08-17T12:44:53Z No. of bitstreams: 1 CECÍLIA ELISA DE SOUSA MUNIZ - DISSERTAÇÃO (PPGEQ) 2017.pdf: 1566496 bytes, checksum: 6a11f4784c5c418c014738b1b8624c76 (MD5) / Made available in DSpace on 2018-08-17T12:44:53Z (GMT). No. of bitstreams: 1 CECÍLIA ELISA DE SOUSA MUNIZ - DISSERTAÇÃO (PPGEQ) 2017.pdf: 1566496 bytes, checksum: 6a11f4784c5c418c014738b1b8624c76 (MD5) Previous issue date: 2017-10-05 / Com os produtos tradicionais fontes de proteína se tornando mais caros a cada dia, surge a necessidade de desenvolver novos produtos alimentícios com alto valor nutricional e que ainda assim sejam de baixo custo, podendo ser consumido pela população de todas as classes econômicas. As barras de cereais já representam uma alternativa de complemento alimentar à base de carboidratos e fibras, sendo assim, esta pesquisa teve como objetivo elaborar barras de cereais que contenham na sua composição resíduos agroindustriais, que submetidos ao processo de enriquecimento proteico por via microbiana, irão aumentar o teor proteico e nutricional deste respectivo produto, além de torna-los mais baratos. O microrganismo utilizado foi a levedura Saccharomyces cerevisiae, e os resíduos escolhidos para tal objetivo foram o bagaço do caju e as cascas de goiabas. Inicialmente fez-se o preparo dos resíduos, os quais foram secos em estufa com circulação de ar a 55±2°C até peso constante. A caracterização físico-química dos resíduos do caju e da goiaba mostrou que, estes poderiam ser submetidos ao processo fermentativo de enriquecimento proteico e posteriormente como ingredientes na elaboração das barras de cereais, apresentando teores suficiente de açúcares redutores, pectina e fibras, além de um pH adequado ao meio de cultivo. Para o enriquecimento proteico utilizou-se a levedura em uma concentração de 3%, para o resíduo do caju e de 5% para o resíduo da goiaba, sob temperatura de 30ºC durante 9h de cultivo. Tanto o resíduo do caju quanto o da goiaba teve o seu teor de proteína elevado em quatro vezes. Após a obtenção dos resíduos enriquecidos proteicamente foram elaboradas quatro formulações de barras de cereais, onde a aveia foi substituída parcialmente pelos resíduos agroindustriais, (F0) formulação de referência sem adição de nenhum dos resíduos, (F1) constituída apenas pelo resíduo do caju, (F2) constituída apenas pelo resíduo da goiaba e (F3) constituída pelos dois resíduos, goiaba e caju. As barras de cereais elaboradas foram caracterizadas físico-químicamente quanto ao teor de água, cinzas, lipídeos, carboidratos, proteína e fibra alimentar, apresentando resultados bastante satisfatórios, principalmente quanto ao teor de proteína, em torno de 12% para a formulação F1, valor este superior ao encontrado nas barras de cereais comercializadas no mercado. Posteriormente elas foram submetidas a análise microbiológica para verificar a ausência/presença de coliformes totais, coliformes fecais e salmonela sp., os resultados obtidos apresentaram contagens inferiores ás máximas estipuladas pela RDC n° 12 atendendo portanto, aos requisitos da legislação vigente. Por fim as formulações foram avaliadas sensorialmente em uma academia da cidade por um grupo de 40 julgadores os quais avaliaram os atributos: cor, aroma, sabor, textura e aceitação global, utilizando uma escala hedônica de 9 pontos, além da intenção de compra, utilizando uma escala hedônica de 5 pontos. Os resultados obtidos foram satisfatórios, para todas as barras de cereais analisadas pelos julgadores os quais afirmaram que gostaram moderadamente (7) e que possivelmente comprariam (4) caso elas fossem comercializadas. / With traditional sources of protein becoming more expensive every day, there is a need to develop new food products with high nutritional value that are still low cost and can be consumed by the population of all economic classes. The cereal bars already represent an alternative food complement based on carbohydrates and fibers, so, this research aims to elaborate cereal bars containing in their composition agroindustrial residues, which undergo the process of microbial protein enrichment, will increase the protein and nutritional content of this respective product, besides making them cheaper. The microorganism used was the yeast Saccharomyces cerevisiae, and the residues chosen for this purpose were the cashew bagasse and the guava husks. Initially the waste was prepared, which were dried in an oven with air circulation at 55 ± 2 ° C until constant weight. The physico-chemical characterization of the cashew and guava residues showed that they could be submitted to the fermentative process of protein enrichment and later as ingredients in the elaboration of the cereal bars, presenting sufficient levels of reducing sugars, pectin and fibers, besides a pH suitable for culture medium. For protein enrichment, the yeast was used at a concentration of 3% for the cashew residue and 5% for the guava residue, at a temperature of 30 ° C for 9h of culture. Both the cashew and guava residue had their protein content elevated fourfold. After obtaining the protein enriched residues, four formulations of cereal bars were elaborated, where the oats were replaced partially by the agroindustrial residues, (F0) reference formulation without addition of none of the residues, (F1) constituted only by the cashew residue, F2) constituted only by guava residue and (F3) constituted by the two residues, guava and cashew. The elaborated cereal bars were physicochemically characterized as water, ash, lipids, carbohydrates, protein and dietary fiber, presenting satisfactory results, mainly regarding the protein content, around 12% for the formulation F1, value higher than that found in cereal bars marketed on the market. Subsequently, they were submitted to microbiological analysis to verify the absence/presence of total coliforms, fecal coliforms and salmonella sp., the results obtained were lower than the maximum stipulated by RDC n° 12, thus meeting the requirements of current legislation. Finally, the formulations were evaluated sensorially in a gymnasium of the city by a group of 40 judges who evaluated the attributes: color, aroma, flavor, texture and global acceptance, using a hedonic scale of 9 points, besides the intention to buy, using a hedonic scale of 5 points. The results obtained were satisfactory for all cereal bars analyzed by the judges who stated that they liked moderately (7) and that they would possibly buy (4) if they were marketed.

The breast cancer susceptibility protein, BRCA1, is a tumor suppressor that helps maintain genomic integrity. Changes in BRCA1 that effect DNA repair processes can fuel cancer induction. The Kelly lab, at the Virginia Tech Carilion Research Institute, has recently developed a new methodology that employs silicon nitride (SiN) microchips to isolate BRCA1 assemblies from the nuclear material of breast cancer cells. These microchips are coated with adaptor proteins that include antibodies against target proteins of interest. The adaptor proteins are added in sequential steps to the coated microchips, followed by an aliquot of sample containing the protein of interest, such as BRCA1. The Kelly lab, partnered with Protochips Inc., developed these devices as a robust, tunable platform to monitor molecular processes, and refer to them as 'Cryo-SiN' in cryo-Electron Microscopy (EM) imaging. We are currently using Cryo-SiN to recruit BRCA1 protein assemblies to the microchip surface under mild conditions, while simultaneously preparing them for cryogenic preservation and EM imaging. This strategy presents a viable alternative to antibody affinity columns that require stringent elution steps to obtain protein complexes from the column. Another advantage of the microchip strategy is that it requires only a 30-minute nuclear extraction, a 60-minute enrichment procedure, and a 5-minute microchip capture step--a total of 95 minutes from initially lysing the cells to plunge-freezing the EM specimens. Therefore, these novel approaches represent a major departure from classical separation procedures that often require days to complete, during which time active protein assemblies can readily dissociate or become inactive. Overall, our use of BRCA1-specific microchips may reveal changes in the BRCA1 architecture during various stages of cancer progression--a major gap in knowledge that persists in cancer research. / M. S.

Submitted by Johnny Rodrigues (johnnyrodrigues@ufcg.edu.br) on 2018-06-05T16:12:57Z No. of bitstreams: 1 GERALDO NEVES OLIVEIRA JÚNIOR - DISSERTAÇÃO PPGSA PROFISSIONAL 2015..pdf: 1634063 bytes, checksum: 80e229d2b1f56448783db163d680a86e (MD5) / Made available in DSpace on 2018-06-05T16:12:57Z (GMT). No. of bitstreams: 1 GERALDO NEVES OLIVEIRA JÚNIOR - DISSERTAÇÃO PPGSA PROFISSIONAL 2015..pdf: 1634063 bytes, checksum: 80e229d2b1f56448783db163d680a86e (MD5) Previous issue date: 2015-12-18