RESUMO
Polyclonal antibodies raised in Balb-c mice against BnSP-7, a Lys-49 phospholipase A2, were used to measure cross reactivity against other snake venoms. Using ELISA, these antibodies were able to recognize PLA2s isoforms present in venoms of botropic snakes at 1:6400, 1:3200 and 1:100 ratios (w/w). These antibodies highly recognized proteins of low molecular weight (â¼14,000) from crude snake venom Bp and Bm by Western Blotting. PLA2 these venoms, by alignment of primary structures demonstrated high identity with BnSP-7 PLA2, especially in the C-terminal region. However, the crude snake venom Bd and Bj, showed low recognition. The PLA2 activity of Bothrops pauloensis, Bothrops moojeni venoms or BpPLA2-TXI was inhibited significantly when anti-BnSP-7 antibodies were incubated at 1:10 and 1:20 ratios (venoms or toxin:anti-BnSP-7, w/w), respectively. The myotoxic effect induced by the same venoms was also reduced significantly at 1:1, 1:10 and 1:20 ratios, by decreased creatine kinase levels. The anti-PLA2 polyclonal antibodies effectively recognized PLA2s from Bothrops pauloensis and Bothrops moojeni venoms, and neutralized specific catalytic and myotoxic activity.
Assuntos
Anticorpos Monoclonais/imunologia , Bothrops/imunologia , Reações Cruzadas/imunologia , Venenos de Crotalídeos/imunologia , Fosfolipases A2/imunologia , Venenos de Serpentes/imunologia , Sequência de Aminoácidos , Animais , Western Blotting , Bothrops/classificação , Bothrops/metabolismo , Venenos de Crotalídeos/metabolismo , Ensaio de Imunoadsorção Enzimática , Masculino , Camundongos Endogâmicos BALB C , Testes de Neutralização , Fosfolipases A2/genética , Fosfolipases A2/metabolismo , Homologia de Sequência de Aminoácidos , Venenos de Serpentes/metabolismo , Especificidade da EspécieRESUMO
Snake venom serine proteases (SVSPs) are enzymes capable of interfering at several points of hemostasis. Some serine proteases present thrombin-like activity, which makes them targets for the development of therapeutics agents in the treatment of many hemostatic disorders. In this study, a recombinant thrombin-like serine protease, denominated rBpSP-II, was obtained from cDNA of the Bothrops pauloensis venom gland and was characterized enzymatically and biochemically. The enzyme rBpSP-II showed clotting activity on bovine plasma and proteolytic activity on fibrinogen, cleaving exclusively the Aα chain. The evaluation of rBpSP-II activity on chromogenic substrates demonstrated thrombin-like activity of the enzyme due to its capacity to hydrolyze the thrombin substrate. These characteristics make rBpSP-II an attractive molecule for additional studies. Further research is needed to verify whether rBpSP-II can serve as a template for the synthesis of therapeutic agents to treat hemostatic disorders.
Assuntos
Bothrops , Serina Proteases/química , Venenos de Serpentes/enzimologia , Sequência de Aminoácidos , Animais , Coagulação Sanguínea/efeitos dos fármacos , Bovinos , Fibrinogênio/química , Transtornos Hemostáticos/tratamento farmacológico , Hidrólise/efeitos dos fármacos , Proteínas Recombinantes/química , Trombina/químicaRESUMO
Objetivos: Avaliar a utilização do teste de avidez de IgG anti-toxoplasma gondii (T. gondii) na rotina laboratorial do pré-natal, entre gestantes do município de Araraquara/SP e região, e analisar sua relação com a realização de tratamento para toxoplasmose durante a gestação e com a ocorrência de toxoplasmose congênita. Métodos: De janeiro a novembro de 2005, no Laboratório de Imunologia Clínica e Biologia Molecular do Centro de Referência Diagnóstica do Núcleo de Atendimento à Comunidade da Universidade Auxiliar Integrada à Faculdade de Ciências Farmacêuticas, Campus de Araraquara, UNESP, Araraquara/SP,analizaram-se os resultados dos testes sorológicos para toxoplasmose na rotina do pré-natal em dois grupos de pacientes. O primeiro constituídos por gestantes com IgM anti-T.gondii positiva (n=33). No grupo com IgM positiva foi realizado o teste de avidez para anticorpos IgG anti-T.gondii.
