Estudo de fluxos metabólicos na produção de Cefamicina C por Streptomyces clavuligerus

• Ano de defesa: 2014
• Autor(a) principal: Cavallieri, André Pastrelo [UNESP]
• Tipo de documento: Tese
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Estadual Paulista (Unesp)
• Palavras-chave em Português: Biotecnologia; Metabolismo; Estafilococos; Penicilina; Metabolism
• Link de acesso: http://hdl.handle.net/11449/110839

Cephamycin C is an antibiotic produced by Streptomyces clavuligerus of great interest due to its higher degree of resistance to β-lactamases as compared to other beta- lactam antibiotics. Cephamycin C is produced in small concentrations in nature as with all secondary metabolites. Therefore, investments in improvement of strains and optimization, of culture media and operation of bioreactors are key aspects to increase the production. However, the effectiveness of such strategies may be limited, requiring the use of new approaches. In this aspect, the metabolic engineering is an important field that combines quantification of metabolic fluxes and molecular techniques for improving strains. The study of metabolic fluxes enables one to identify metabolic bottlenecks through careful analysis of metabolism of the microorganism, and to suggest ways to increase production. In this project we carried out the study of metabolic fluxes in S. clavuligerus aiming to find ways to increase the production of cephamycin C. For this, first we developed a chemically defined culture medium because this kind of media does not cause interference in the analyses. The developed medium contained maltose as carbon source and lysine as nitrogen source and resulted in 9 g.L-1 of biomass and 200 mg.L-1 of cephamycin C in the continuous process. This mode of operation was only possible in the bioreactor due to its pH control. Due to variations in the pH, the continuous process in shaken-flasks became unviable. The metabolic model was constructed with 78 reactions and 81 metabolites (10 external and 71 internal) which suitably described the metabolism of S. clavuligerus. This model was simulated with the aid of Optflux, a multi-task software developed for this purpose. The profiles of maltose, lysine, biomass, cephamycin C, clavulanic acid, external protein and CO2 evolution were monitored. These data were used for the model simulations. The results allowed...