Oxidation of monohydric phenol substrates by tyrosinase. An oximetric study

摘要: The purity of commercially available mushroom tyrosinase was investigated by non-denaturing PAGE. Most the protein in preparation migrated as a single band under these conditions. This contained both and dopa oxidase activity. No other activity either classification found preparation. Oxygen consumption during oxidation monohydric phenol substrates tyrosine 4-hydroxyanisole (4HA) monitored oximetry order to determine stoichiometry reactions. For complete oxidation, molar ratio oxygen: 4HA 1:1. Under identical conditions, required 1.5 mol oxygen/mol tyrosine. additional oxygen uptake is due internal cyclization dopaquinone form cyclodopa, which undergoes redox reaction with dopachrome dopa, then oxidized enzyme, leading an 0.5 original substrate. for 200 nmol constant over range concentrations 33-330 units/ml maximum rate directly proportional concentration tyrosinase, whereas length lag phase decreased non-linearly increasing concentration. Activation enzyme exposure citrate not seen, nor abolished low pH. Michaelis-Menten analysis pre-exposure dithiothreitol gave Km values 153 20 microM respectively.