Tyrosinase activity in the uveal tissue of the adult bovine eye

Abstract

Tyrosinase activity in crude extracts from various tissues of the adult bovine eye was examined biochemically. Enzyme activity was measured by using l-3,4-dihydroxyphenylalanine ( l-DOPA) as substrate and determining colorimetrically by an increase in absorbancy at 400 or 475 nm. Tyrosinase activity was found in the ciliary body, iris, and choroid with the ciliary body having the highest enzyme activity. The enzyme was 1324-fold purified from the crude extract of the ciliary body by ammonium sulfate fractionation, trypsin digestion, followed by chromatography on Sephacryl S-200, hydroxylapatite, and DEAE-cellulose columns. The apparent K m value for l-DOPA was 0·2 m m and the molecular weight of the enzyme was estimated to be 70000 by gel method. The enzyme activity was markedly reduced by phenylthiourea and diethyldithiocarbamate, specific inhibitors of tyrosinase.