Isolation and characterization of a tyrosinase from the skin of the white silky fowl (Gallina lanigera) employing copper saturated diethylaminoethyl-cellulose.
Tyrosinase was purified to apparent homogeneity from the skin of the White Silky fowl (Gallina lanigera). Under alkaline conditions copper ions from a complex with the diethylaminoethyl moiety of DEAE-cellulose, allowing the purification of the enzyme in a one step chromatography. In polyacrylamide gel electrophoresis containing sodium dodecyl sulfate the purified tyrosinase displays a single band with a relative molecular mass of 66,000 Da. The crystalline tyrosinase was obtained from 32-33% saturated solution of ammonium sulfate. The enzyme catalyzes the o-hydroxylation of L-tyrosine to L-dihydroxyphenylalanine as well as the oxidation of pyrocatechol to o-benzoquinone. For the pyrocatechol oxidation the Michaelis Menten constant Km, is calculated to be around 1.19 x 10(-3) M.