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Charakterizace N-demetyllinkomycin-metyltransferázy. / Characterization of N-demethyllincomycin-methyltransferase.Poľan, Marek January 2010 (has links)
Lincomycin is a naturally occurring member of a lincosamide group of antibiotics. The cluster of lincomycin biosynthetic gene was already decribed and the function of many of genes has been clarified. This work, "Characterization of N-demethyllincomycin-methyltransferase", is focused on the study of the final step of lincomycin biosynthetic pathway - the methylation of nitrogen atom from the pyrollo ring of the propylproline unit of the N-demethyllicomycin (NDL). The aim of this work was the characterization of the protein LmbJ, catalysing this final biosynthetic step. All the experiments were provided for the enzyme LmbJ with N-terminal histidine tag, which had been prepared by the heterologous expression in E.coli cells. The pH and temperature optimum was determined as well as the Michaelis constants for both substrates of the reaction - N-demethyllincomycin and S-adenosyl methionine (SAM - a methyl group donor). With the exception of the pH optimum, all specified parameters have markedly differed from the data published for the enzyme isolated from the natural source. Based on the comparison of electron microscopy, blue native gel electrophoresis and gel filtration results, the hypothetical model of the LmbJ quarternary structure was created. Majority of methyltranserases, so far described occure in...
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Análise genômica de Streptomyces olindensis DAUFPE 5622 e de suas vias crípticas para a obtenção de novos metabólicos secundários de interesse biotecnológico. / Analysis of Streptomyces olindensis DAUFPE 5622 genome and its cryptic pathways to obtain new secondary metabolites of biotechnological interest.Torres, Maria Alejandra Ferreira 08 December 2015 (has links)
Os compostos de origem microbiana tem readquirido interesse pela biodisponibilidade, especificidade de alvo e diversidade química, mas as vias biosintéticas permanecem crípticas em condições de cultura. Uma estratégia para expressa-las é a super-expressão de genes ativadores. O laboratório de Bio-Produtos no ICB na USP tem trabalhado com Streptomyces olindensis produtor da Cosmomicina D uma molécula com atividade antitumoral de interesse devido ao padrão de glicosilação. O genoma de S. olindensis foi sequenciado e submetido ao NCBI (JJOH00000000) e utilizando o software antiSMASH foram identificados 33 clusters envolvidos na produção de metabolitos secundários. Encontraram-se clusters gênicos para a produção de metabolitos como Melanina, Geosmina, entre outros. Além, foi realizada uma analise de genômica comparativa para caracterizar e anotar as 22 vias biossintéticas desconhecidas em S. olindensis. Finalmente, escolheram-se a via do aminociclitol e um Policetídeo Tipo I para a super-expressão de genes reguladores levando a detecção do composto sob condições de cultura. / Microbial metabolites regain interest due to its bioavailability, target specificity and chemical diversity, but the biosynthetic pathways remain silenced under culture conditions. A strategy to obtain them is the over expression of regulatory genes. Bio-products laboratory at USP has been working with Streptomyces olindensis, products of Cosmomycin D, an antitumoral molecule with a distinctive glycosylation pattern. S. olindensis genome was sequenced and submitted to NCBI (JJOH00000000) and employing antiSMASH server 33 secondary metabolite related clusters were identified. Known pathways were found such as genes for melanin production, Geosmin and others. Additionally, a comparative genomic approach was used to characterize the 22 biosynthetic unknown pathways described in S. olindensis. Subsequently, Aminocyclitol and Polyketide Type I were chosen to evaluated, over expressing the regulatory genes, leading to the compound detection in regular culture conditions.
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Análise genômica de Streptomyces olindensis DAUFPE 5622 e de suas vias crípticas para a obtenção de novos metabólicos secundários de interesse biotecnológico. / Analysis of Streptomyces olindensis DAUFPE 5622 genome and its cryptic pathways to obtain new secondary metabolites of biotechnological interest.Maria Alejandra Ferreira Torres 08 December 2015 (has links)
Os compostos de origem microbiana tem readquirido interesse pela biodisponibilidade, especificidade de alvo e diversidade química, mas as vias biosintéticas permanecem crípticas em condições de cultura. Uma estratégia para expressa-las é a super-expressão de genes ativadores. O laboratório de Bio-Produtos no ICB na USP tem trabalhado com Streptomyces olindensis produtor da Cosmomicina D uma molécula com atividade antitumoral de interesse devido ao padrão de glicosilação. O genoma de S. olindensis foi sequenciado e submetido ao NCBI (JJOH00000000) e utilizando o software antiSMASH foram identificados 33 clusters envolvidos na produção de metabolitos secundários. Encontraram-se clusters gênicos para a produção de metabolitos como Melanina, Geosmina, entre outros. Além, foi realizada uma analise de genômica comparativa para caracterizar e anotar as 22 vias biossintéticas desconhecidas em S. olindensis. Finalmente, escolheram-se a via do aminociclitol e um Policetídeo Tipo I para a super-expressão de genes reguladores levando a detecção do composto sob condições de cultura. / Microbial metabolites regain interest due to its bioavailability, target specificity and chemical diversity, but the biosynthetic pathways remain silenced under culture conditions. A strategy to obtain them is the over expression of regulatory genes. Bio-products laboratory at USP has been working with Streptomyces olindensis, products of Cosmomycin D, an antitumoral molecule with a distinctive glycosylation pattern. S. olindensis genome was sequenced and submitted to NCBI (JJOH00000000) and employing antiSMASH server 33 secondary metabolite related clusters were identified. Known pathways were found such as genes for melanin production, Geosmin and others. Additionally, a comparative genomic approach was used to characterize the 22 biosynthetic unknown pathways described in S. olindensis. Subsequently, Aminocyclitol and Polyketide Type I were chosen to evaluated, over expressing the regulatory genes, leading to the compound detection in regular culture conditions.
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