Estudos estruturais com fosfolipases A2 isoladas de venenos de serpentes dos gêneros Bothrops e Crotalus

• Ano de defesa: 2013
• Autor(a) principal: Fernandes, Carlos Alexandre Henrique [UNESP]
• Tipo de documento: Tese
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Estadual Paulista (Unesp)
• Palavras-chave em Português: Serpente peçonhenta - Genética; Cobra - Veneno; Biologia molecular; Fosfolipases; Molecular biology
• Link de acesso: http://hdl.handle.net/11449/108406

Snakebite is a very serious public health problem still today, affecting tropical and subtropical countries, especially the rural areas of Africa, Asia, Oceania and Latin America. In Brazil, Bothrops and Crotalus snake genus are responsible of almost 90% of snakebites. The venoms of Bothrops snakes have a class of phospholipases A2, known as Lys49-PLA2s, which do not have catalytic activity. However, they are able to induce myonecrosis by a non-catalytic mechanism that is not fully understood. In the venom of Crotalus snakes, a phospholipase A2, known as crotoxin constitutes about 60% of the venom from these animals. This protein is a potent neurotoxin that causes a large block of neuromuscular transmission formed by complexation of crotoxin A (non-catalytic) and crotoxin B (catalytic). In this work, it was used several theoretical and experimental techniques, in particular X-ray protein crystallography, to generate information to extend the knowledge of the action of these proteins. Regarding to crotoxin, the crystal structure of crotoxin B isolated from Crotalus durissus colillinetaus was solved and the heterologous expression of isoform CBa2 of crotoxin B from Crotalus durissus terrificus was performed, in order to, subsequently, produce site-directed mutants. Furthermore, it is presented small angle X-ray scattering experiments with the crotoxin and its isolated subunits that provided intriguing and novel information about the oligomerization of these proteins. Regarding to Lys49-PLA2s from Bothrops snakes, it is presented two new crystal structures of this class isolated from Amazonian snake B. brazili. The analysis of these structures and the structural comparison to other Lys49-PLA2s crystallographic structures generated new information about the position of hydrophobic residues in C-terminal portion of these proteins. This information led us to a new proposition of the mechanism of action of these proteins. It is also presented ...