TYROSINASE-CATALYZED BINDING OF 3,4-DIHYDROXYPHENYLALANINE WITH PROTEINS THROUGH THE SULFHYDRYL-GROUP

The cytotoxicity of catechols has been ascribed to covalent binding of the o-quinone oxidation products to proteins through sulfhydryl groups. The nature of the covalent binding was studied with dopaquinone formed on tyrosinase oxidation of 3,4-dihydroxyphenylalanine (DOPA). After acid hydrolysis of the reaction products, cysteinyldopas liberated (protein-bound cysteinyldopas) were determined by HPLC with electrochemical detection. When 0.1 mM DOPA was oxidized in the presence of 0.2 mM bovine serum albumin, alcohol dehydrogenase or isocitrate dehydrogenase, protein-bound cysteinyldopas were formed in yields of 5.4, 44, or 33%, respectively. The covalent binding was almost completely inhibited by 1 mM cysteine or 1 mM ascorbic acid, but 10 mM lysine had no effect. These results unambiguously demonstrate that dopaquinone can bind with proteins mostly through sulfhydryl groups.