Evidence for covalent lipoyl adduction with dopaquinone following tyrosinase-catalyzed oxidation

Previous studies have examined the conjugation of sulfhydryl compounds such as l-cysteine and glutathione with DOPA-quinone following the oxidation of tyrosine and DOPA by tyrosinase. These covalent reactions play a key role in the regulation and metabolism of pigment cells. We report on the first direct evidence for the formation of lipoyl adducts in reactions of thiol groups with DOPA-quinone in dihydrolipoic acid (6,8-dimercaptooctanoic acid [DHLA]). Incubating DHLA with DOPA-quinone followed by tyrosinase-catalyzed oxidation resulted in the three products predicted by HPLC-UV and LC-ESI−-MS analyses for DHLA DOPA conjugates. In the current study, we identified 5-S-lipoyl-DOPA among the principal products isolated by HPLC and characterized by FAB−-MS, ESI−-MS/MS, and 1H NMR, 2D-COSY studies. Collectively, these results suggest that DHLA undergoes sulfhydryl conjugation with DOPA-quinone, pointing to the involvement of thiol-reactive metabolites.