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Generating an expression construct and soluble protein for characterization studies of a putative RNA m5C methyltransferase, yeast ORF YNLO22cCraft, Jennifer Leigh January 2005 (has links)
RNA m5C methyltransferases are a group of enzymes that catalyze the transfer of a methyl group to a cytosine nucleotide of RNA. Only two of these enzymes have been well characterized: Fmu from E. coli and Trm4p from S. cerevisiae. YNLO22c is one of three ORFs identified in S. cerevisiae that have homology with both known and putative RNA m5C methyltransferases, but its encoded protein, YNLO22p, has not been confirmed to have enzyme activity. Verifying that YNLO22c encodes an RNA m5C methyltransferase will require adequate amounts of soluble YNLO22p for enzyme assays. A bacterial expression plasmid for YNLO22c was developed, but the result was insoluble protein. Therefore, several methods known to improve protein solubility were tested to develop a system in which a sufficient amount of soluble YNLO22p could be produced. Results of this study found that coexpression of YNLO22c with chaperone proteins can provide sufficient quantities of soluble YNLO22p. / Department of Biology
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Extração, concentração e caracterização fisico-quimica e funcional das proteinas de semente de cupuaçu (Theobroma grandiflorum Schum) / Extration, concentration and functionary and physical-chemical characterization of proteins of cupuacu seed(Theobroma grandiflorum Schum)Carvalho, Ana Vania 24 March 2004 (has links)
Orientador: Nelson Horacio Pezoa Garcia / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-03T20:56:29Z (GMT). No. of bitstreams: 1
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Previous issue date: 2004 / Resumo: O cupuaçu (Theobroma grandiflorum Schum) é uma fruta típica da região Norte do Brasil, com grande potencial econômico. Atualmente é a polpa que sustenta a produção, industrialização e comercialização desta fruta. A semente, um subproduto da industrialização da polpa, começa a despertar interesse como um produto de alto valor nutricional e com grande potencial de mercado para produtos industrializados. O presente estudo teve como objetivo a produção e caracterização de farinha, concentrado e isolado protéico da semente do cupuaçu, quanto a sua composição protéica e de aminoácidos e características químicas e funcionais. A farinha, obtida das sementes trituradas e desengorduradas, serviu como matéria-prima para a extração da fração protéica em meio aquoso, obtendo-se o concentrado protéico (precipitação em pH 3,5) e o isolado protéico (solubilização em pH 9 e precipitação em pH 3,5). A farinha desengordurada, o concentrado e o isolado protéico de semente de cupuaçu foram submetidos à análise de composição centesimal, cor, aminoácidos, perfil eletroforético de suas proteínas e propriedades funcionais. O concentrado apresentou um teor protéico de 31,18% e o isolado de 64,33%. Quanto ao perfil eletroforético de suas proteínas, observou-se a presença de três principais bandas protéicas, com pesos moleculares variando de 20,03 a 39,79kDa além de três bandas fracas adicionais. As bandas mais fracas parecem ter sido seletivamente perdidas durante a extração alcalina para produção do isolado protéico, estando presentes apenas na farinha desengordurada e no concentrado protéico. Observou-se que a farinha desengordurada, o concentrado e o isolado protéico de semente de cupuaçu, apresentaram boa composição aminoacídica, com teores superiores aos recomendados para a grande maioria dos aminoácidos. Tanto a farinha como o concentrado apresentaram muito boa capacidade de retenção de água e de óleo. Observou-se também, boa capacidade emulsificante em pH 7,0, para a farinha, o concentrado e o isolado protéico, 987,50, 977,50 e 1380,00mL óleo/g produto, respectivamente. Assim, estes produtos apresentam potencial de utilização, não somente para enriquecer outros alimentos, mas também para melhorar certas propriedades funcionais / Abstract: Cupuacu (Theobroma grandiflorum Schum) is a native fruit from northern Brazil, with great economic potential. Its productive chain is currently sustained by the pulp market. The seed, treated as a by-product of the pulp industrialization, has recently come to public attention as a highly nutritive product with great market potential for industrialized products. The objective of the present study was the production and characterization of flour, protein concentrate and protein isolate obtained from the cupuacu seed, in terms of their chemical and functional properties and their protein and amino acid composition. The flour, obtained from ground and defatted seeds, was used to extract the protein fraction in an aqueous medium, obtaining the protein concentrate (precipited at pH 3.5) and the protein isolate (solubilized at pH 9.0 and precipited at pH 3.5). The defatted flour, the protein concentrate and the protein isolate were analyzed in terms of their chemical composition, color, amino acid contents, electrophoretic protein profile and functional properties. The concentrate presented a protein content of 31.18% and the isolate, 64.29%. The electrophoretic profile of their proteins indicated the presence of three main protein bands, with molecular weights ranging from 20.03 to 39.79 kDa, besides three additional weak bands. The weaker bands seem to have been selectively lost during alkaline extraction for production of the protein isolate, as they were detected only in the defatted flour and in the protein concentrate. The defatted flour, the protein concentrate and the protein isolate obtained from cupuacu presented a good amino acid composition, as the contents of most amino acids were higher than the daily recommended amounts. The flour and the concentrate presented both excellent water and oil retention capacity. Good emulsifying ability at pH 7.0 was also observed for all three products: 987.50 mL oil/g for the flour, 977.50 mL oil/g for the protein concentrate and 1380.00 mL oil/g for the protein isolate. So, these products presented good utilization potential, not only to enrich other foods, but also to enhance some functional properties / Doutorado / Doutor em Tecnologia de Alimentos
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