Characterization of phenoloxidase activity in Sydney rock oysters (Saccostrea glomerata).
Phenoloxidase (PO) activity was studied in Sydney rock oysters (Saccostrea glomerata). As in other molluscs, PO was found to exist as a pro-enzyme (proPO) in hemocytes. ProPO could be activated to PO by exogenous proteases (trypsin and chymotrypsin), exposure of hemocytes to pathogen-associated molecular patterns (PAMPs) and by the detergents, Triton X-100 and sodium dodecyl sulphate (SDS). Inhibition studies confirmed the proPO activating system of Sydney rock oysters is a proteinase cascade in which Ca2+ dependent serine proteinases proteolytically convert proPO into active PO. Activated PO was found to be a tyrosinase-like enzyme that is responsible for both monophenolase and diphenolase activity. The bifunctional PO had higher affinity for the monophenol, hydroquinine monomethyl ether (4HA) (Km=4.45+/-1.46 mM) than for the diphenol, l-DOPA (Km=10.27+/-1.33 mM). Maximum enzyme activity was evident at 37 degrees C, pH 8 and at salinities of between 30 and 37 ppt. Melanogenesis catalysed by the active enzyme is a composite of eumelanin and the product of a sclerotin pathway combining DOPA decarboxylase with PO activity.
Aladaileh
Saleem
S
Department of Biological Sciences, Macquarie University, North Ryde, NSW, 2109, Australia.
Rodney
Peters
P
Nair
Sham V
SV
Raftos
David A
DA
D000818:Animals; D003902:Detergents / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000378:metabolism; D006434:Hemocytes / Q000201:enzymology; D006458:Hemolymph / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D019746:Intramolecular Oxidoreductases / Q000378:metabolism; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology; D014442:Monophenol Monooxygenase / Q000037:antagonists & inhibitors / Q000097:blood / Q000378:metabolism*; D010125:Ostreidae / Q000201:enzymology*; D011480:Protease Inhibitors / Q000494:pharmacology; D011802:Quinidine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D054712:Salinity; D012697:Serine Endopeptidases / Q000378:metabolism; D013696:Temperature; D015054:Zymosan / Q000378:metabolism / Q000494:pharmacology
D003902:Detergents; D004792:Enzyme Precursors; D008070:Lipopolysaccharides; D011480:Protease Inhibitors; C014486:hydroquinidine; D015054:Zymosan; D014442:Monophenol Monooxygenase; D012697:Serine Endopeptidases; D019746:Intramolecular Oxidoreductases; C042501:dopachrome isomerase; D011802:Quinidine