PURIFICATION AND PROPERTIES OF TYROSINASE ISOZYMES FROM THE GILL OF LENTINUS-EDODES FRUITING BODY

Six tyrosinase isozymes were purified from the browned gill of the fru iting body of Lentinus edodes by ammonium sulfate fractionation, DEAE- Sephacel and Q-Sepharose column chromatography, and partially denaturi ng SDS-PAGE. At the step of Q-Sepharose column chromatography, two act ive fractions (A and B) were obtained. Each fraction was separated to three further fractions, A1, A2, and A3, and B1, B2, and B3, respectiv ely, by partially denaturing SDS-PAGE. All these isozymes consisted of two types of polypeptides: alpha polypeptide (A alpha or B alpha) and either beta (A beta or B beta) or gamma polypeptide (A gamma or B gam ma). The alpha polypeptide contained the consensus amino acid sequence of the active site of known tyrosinases, which is considered to act a s a catalytic subunit. From the results of peptide mapping and the ami no acid composition, A alpha and B alpha polypeptides mere considered to be different proteins. The kinetic properties of the purified tyros inase isozymes differed greatly according to whether they contained be ta or gamma polypeptide, indicating these polypeptides to be a possibl e regulatory subunit.