Ditopic Chelators of Dicopper Centers for an Enhancement of Tyrosinases Inhibition.

Tyrosinase enzymes (Tys) are involved in the key steps of melanin (protective pigments) biosynthesis and molecules targeting the binuclear copper active site on tyrosinases represent a relevant strategy to regulate enzyme activities. In this work, we studied the possible synergic effect generated by combination of known inhibitors. For this, derivatives containing kojic acid (KA) and 2-Hydroxypyridine- N -oxide (HOPNO) combined with a thiosemicarbazone (TSC) moiety were synthetized. Their inhibition activities were evaluated on purified tyrosinases from different sources (mushroom, bacterial and human) as well as on melanin production by lysats from human melanoma MNT-1 cell line. Results showed significant enhancement of the inhibitory effects compared to the parent compounds, in particular for HOPNO-TSC. In order to elucidate the interaction mode with the dicopper(II) active site, we investigated the binding studies towards a tyrosinase bio-inspired model of the dicopper(II) center. The structure of the isolated adduct between one ditopic inhibitor (KA-TSC) and the model complex reveals that the binding to a dicopper center can occur with both chelating sites. Computational studies on model complexes and docking studies on enzymes led to the identification of KA and HOPNO moieties as interacting groups with the dicopper active site.