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MARTINS, L. M. Effect of an anti-apoptotic protein obtained from the hemolymph of Lonomia obliqua on the mitochondria of cells Sf-9. 2011. 87 p. Master thesis (Biotecnology) Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, 2011. The protein expression system using baculovirus as a vector has been intensively used for the production of recombinant proteins because it allows the formation of posttranslational modifications and has the presence of a strong promoter, polyhedrin. However, in some cases, this expression system is not as effective because the infection of cells by baculovirus induces death by apoptosis, thereby limiting the industrial production of various recombinant proteins of interest. Mitochondria have an important role in controlling the process of death by apoptosis, because during the transduction of a signal of apoptosis in a cell, there is a change in membrane permeability of mitochondria, resulting in the formation of pores, which can be caused by loss of mitochondrial membrane electrochemical potential (m). The formation of these pores leads to the release of apoptogênicas proteins such as cytochrome c in the cytosol, which consequently activate proteolytic proteins known as caspases and DNase and the dying process becomes irreversible. We report the occurrence of apoptosis in cultured insect cells and mammalian depleted of nutrients or induced by chemical and viral agents, and supplementation of these cultures with hemolymph of Lonomia obliqua is able to extend the viability of the culture avoiding death by apoptosis. This study aimed to determine in advance the anti-apoptotic mechanism of action of a protein isolated from hemolymph of Lonomia obliqua. The identification of this mechanism may help develop strategies for controlling apoptosis in cell culture allowing the optimization of productivity of viral and recombinant proteins. Tests for antiapoptotic activity of crude hemolymph were performed in Sf-9 cells and were found that supplementation of cell cultures with only 1% of hemolymph was sufficient to enhance growth and viability of cells for at least three days. The brute and the fraction of the hemolymph protein isolate by liquid chromatography were used in experiments aimed at determining the effect of protein on the mitochondria, checking its action on the mitochondrial membrane potential. The data from this study suggest that this protein is the action on the membranes of mitochondria, being able to maintain high mitochondrial membrane potential, since in all experiments, there was a death by blocking apoptosis, with clear protection mitochondrial membrane potential.

Estudo do efeito de uma proteína antiapoptótica obtida da hemolinfa de Lonomia obliqua sobre as mitocôndrias de células Sf-9.