Tyrosinases from two different loci are expressed by normal and by transformed melanocytes

Two pigmentation related genes have recently been cloned which map to the brown (b) and albino (c) loci of mice; these loci influence the quality and quantity, respectively, of melanin produced by melanocytes. Both these gene products are biochemically similar and have extensive amino acid sequence similarity to each other and to lower forms of tyrosinase (EC 1.14.18.1), a copper binding enzyme responsible for melanin production. In order to characterize the catalytic activities of these molecules, we have synthesized peptides and prepared antibodies to them which specifically recognize the gene products in question. By use of immune affinity purification protocols, we have isolated the proteins encoded by the brown and albino loci and have determined that both have the catalytic functions ascribed to tyrosinase, i.e. hydroxylation of tyrosine to 3,4-dihydroxyphenylalanine (DOPA) and the oxidation of DOPA to DOPAquinone. These are the critical reactions to melanogenesis since melanin pigment can be spontaneously produced from those products. The specific activity of the albino locus encoded product is considerably higher than that of the protein encoded by the brown locus, although the latter protein is present in higher quantity in melanocytes than is the protein encoded by the albino locus. These results are surprising since it was anticipated that tyrosinase was the product of single gene locus, and suggest that regulation of melanogenesis in mammals is controlled at the enzymatic level by several different gene products.