Estudo da função biológica da oxidase alternativa (AOX) de Moniliophthora perniciosa (fungo da vassoura de bruxa) em Saccharomyces cerevisiaedissertation
Аннотация: Moretti-Almeida, G. Study of the biological function of the Alternative Oxidase (AOX) from Moniliophthora pernicious (witches' broom fungus) in Saccharomyces cerevisiae . 80p. Thesis (Master) Dissertacao (Mestrado) – Faculdade de Ciencias Farmaceuticas, Universidade de Sao Paulo. Sao Paulo, 2014. Moniliophthora pernicious is a basidiomycete fungus that causes witches' broom of cacao disease. A number of techniques for disease control has been applied and tested, including the use of the fungicides that inhibits the main respiratory chain, specific to fungi. However, M. pernicious has proven to be resistant to these drugs and one possible explanation to this resistance would be the activity of an alternative oxidase activity (AOXp), whose expression and activity have already been characterized. Functional analysis of this gene in M. pernicious was not performed because an effective technique for producing mutants of the fungus has not yet been established. This project aimed to test the hypothesis that the expression of Mp-AOX gene prevents oxidative stress due to decreased production of reactive oxygen species (ROS) by the respiratory chain. To perform this characterization, MpAOX gene was cloned and heterologous expressed in Saccharomyces cerevisiae. The yeast expression vector pYES2/CT with Mp-AOX gene was constructed and the yeast strain W303 1b transformed. Analyses measuring cell mass formation, generation of mitochondrial reactive oxygen species (ROS), viability tests challenging the cells against inhibitors of the main respiratory chain, Antimycin A and azoxystrobin, and against the specific inhibitor of the AOXp salicylaldehyde hydroxamic acid (SHAM) were performed. Our results indicate that the Mp-AOX gene expression in S. cerevisiae causes decreased generation of cellular biomass, less cell proliferation and the decay in the production of ROS, suggesting that our initial hypothesis that Mp-AOXp reliefs the oxidative stress is correct. The recombinant yeast expressing MpAOX gene was feasible for use as a model for studies of new chemical to fight witches' broom, helping with agriculture and promoting new understandings to combat this fungus.
Год издания: 2014
Авторы: Gabriel Moretti de Almeida
Ключевые слова: Fungal and yeast genetics research, Enzyme-mediated dye degradation, Fermentation and Sensory Analysis
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