Characterization of Phenoloxidase Activity From Spider <i>Polybetes pythagoricus</i> Hemocyanin
Hemocyanin of the spider <i>Polybetes pythagoricus</i>, in addition to its typical role as an oxygen transporter, also exhibits a phenoloxidase activity induced by micellar concentrations of SDS. In the present work, we found the kinetic parameters K<sub>m</sub> and V<sub&...
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| Autores principales: | , , , , |
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| Formato: | Articulo Preprint |
| Lenguaje: | Inglés |
| Publicado: |
2015
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| Materias: | |
| Acceso en línea: | http://sedici.unlp.edu.ar/handle/10915/127717 |
| Aporte de: |
| Sumario: | Hemocyanin of the spider <i>Polybetes pythagoricus</i>, in addition to its typical role as an oxygen transporter, also exhibits a phenoloxidase activity induced by micellar concentrations of SDS. In the present work, we found the kinetic parameters K<sub>m</sub> and V<sub>max</sub> of <i>Polybetes pythagoricus</i> hemocyanin (<i>Pp</i>Hc) PO activity to be 0.407mM and 0.081 μmolmin⁻¹mg protein⁻¹, respectively. Dopamine was used as the substrate with SDS at a final concentration of 10mM and a 30-min incubation at 25°C. Conformational changes in Hc associated with the SDS treatment were analyzed using far-UV circular dichroism, intrinsic fluorescence and absorption spectroscopy. The secondary and tertiary structural changes of <i>Pp</i>Hc induced by SDS led to increases in α-helical content and tryptophan fluorescence intensity. A reduction in the absorption spectrum at 340 nm in the presence of SDS was also observed. These results suggest that the SDS-induced PO activity of <i>Pp</i>Hc can be ascribed to conformational changes in the local environment of the typer-3 copper active site. |
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