Purification and characterization of a phosphodiesterase from Bothrops alternatus snake venom |
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Authors: | Valério A A Corradini A C Panunto P C Mello S M Hyslop S |
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Institution: | (1) Departamento de Farmacologia, Faculdade de Ciências Médicas, Universidade Estadual de Campinas (UNICAMP), CP 6111, 13083–970 Campinas, SP, Brazil;(2) Centro de Controle de IntoxicaÇões, Hospital das Clínicas, Universidade Estadual de Campinas (UNICAMP), CP 6111, 13083–970 Campinas, SP, Brazil |
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Abstract: | A phosphodiesterase was purified from the venom of the snake Bothrops alternatus by a combination of gel filtration and ion exchange chromatographies. In SDS-PAGE, the enzyme gave a single band with a molecular mass of 105 kDa, which was unaltered in the presence of -mercaptoethanol, indicating that the protein contained no subunits. A single protein band was also observed in native PAGE. There were no contaminating 59-nucleotidase, alkaline phosphatase and protease activities. The enzyme was recognized by commercial bothropic antiserum and gave a single band in immunoblotting. The enzyme had a pH optimum in the range of 7.5–9.5 and the optimum temperature was 60°C, with activity being rapidly lost within 1 min at 70°C. The Km of the enzyme was 2.69 mM. PDE activity was potentiated by cobalt and, to a lesser extent, by calcium, whereas copper, manganese, zinc, EDTA, and -mercaptoethanol were inhibitory. These properties show that this enzyme is very similar to that isolated from other snake venoms. |
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Keywords: | Bothrops alternatus enzyme phosphodiesterase purification venom |
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