Characterization of Phenoloxidase Activity From Spider Polybetes pythagoricus Hemocyanin

LAINO ALDANA; LAVARÍAS SABRINA; SUAREZ GUSTAVO; LINO AGUSTINA; CUNNINGHAM MÓNICA

JOURNAL OF EXPERIMENTAL ZOOLOGY PART A-ECOLOGICAL GENETICS AND PHYSIOLOGY

WILEY-LISS, DIV JOHN WILEY & SONS INC

Lugar: New York; Año: 2015 vol. 323 p. 547 - 555

1932-5223

Hemocyanin of the spiderPolybetes pythagoricus, 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 KmandVmaxofPolybetes pythagoricus hemocyanin (PpHc) PO activity to be 0.407 mM and 0.081mmolmin1 mg protein1, respectively.Dopamine was used as the substrate with SDS at afinal concentration of 10 mM and a 30-min incubation at 25°C. Conformational changes in Hc associated with the SDS treatment wereanalyzed using far-UV circular dichroism, intrinsicfluorescence and absorption spectroscopy. The secondary and tertiary structural changes ofPpHc induced by SDS led to increases ina-helical content and tryptophanfluorescence 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 PpHc can be ascribed to conformational changes in the local environment of the typer-3 copper active site.