Melanization and Tyrosinase Activity

Purified soluble tyrosinase isolated from the melanosomes of Harding-Passey mouse melanoma was incubated with L-dopa. The reaction velocity of the tyrosinase thus incubated decreased significantly. There was an inverse linear relationship between the concentration of dopa in the preincubation mixture and the reaction velocity of the tyrosinase which remained. The experimental results obtained were similar to what had been observed in melanosomes.64Cu-exchanged tyrosinase or 11C-amino-acid-labeled tyrosinase were incubated with L-dopa; the reaction mixture was centrifuged after varied incubation times, and the radioactivity of the obtained supernatant was determined. The time courses of the disappearance of labeled tyrosinase from the reaction systems indicated that soluble tyrosinase combined with the products of the dopa-tyrosinase reaction and precipitated out with them. Thus tyrosinase was excluded from the reaction systems.The experimental results obtained suggest that the decrease of tyrosinase activity is due to the binding of tyrosinase with reaction products, probably at the active site, and to the disappearance of the tyrosinase molecule from the reaction system