New Inhibitors of Laccase and Tyrosinase by Examination of Cross-Inhibition between Copper-Containing Enzymes

Abstract

Coppers play crucial roles in the maintenance homeostasis in living species. Approximately 20 enzyme families of eukaryotes and prokaryotes are known to utilize copper atoms for catalytic activities. However, small-molecule inhibitors directly targeting catalytic centers are rare, except for those that act against tyrosinase and dopamine-β-hydroxylase (DBH). This study tested whether known tyrosinase inhibitors can inhibit the copper-containing enzymes, ceruloplasmin, DBH, and laccase. While most small molecules minimally reduced the activities of ceruloplasmin and DBH, aside from known inhibitors, 5 of 28 tested molecules significantly inhibited the function of laccase, with the Ki values in the range of 15 to 48 µM. Enzyme inhibitory kinetics classified the molecules as competitive inhibitors, whereas differential scanning fluorimetry and fluorescence quenching supported direct bindings. To the best of our knowledge, this is the first report on organic small-molecule inhibitors for laccase. Comparison of tyrosinase and DBH inhibitors using cheminformatics predicted that the presence of thione moiety would suffice to inhibit tyrosinase. Enzyme assays confirmed this prediction, leading to the discovery of two new dual tyrosinase and DBH inhibitors.

Some Antioxidant Properties of Components from the Flower of Ochna integerrima and Their Beneficial Effects on HaCaT Keratinocytes and in Silico Analysis on Tyrosinase

“…These results are clearly a favorable premise that allows further molecular modeling studies to determine the mechanism of oxidoreductase enzyme inhibition. Specifically, tyrosinase (Ty), a copper‐containing monooxygenase responsible for catalyzing melanin synthesis, is a strong candidate [18] . Reportedly, phenolic secondary metabolites could inhibit enzymatic activity through hydrogen bond formation of hydrogen groups with the active site of the tyrosinase enzyme.…”

“…Specifically, tyrosinase (Ty), a copper-containing monooxygenase responsible for catalyzing melanin synthesis, is a strong candidate. [18] Reportedly, phenolic secondary metabolites could inhibit enzymatic activity through hydrogen bond formation of hydrogen groups with the active site of the tyrosinase enzyme. Molecular docking simulations are a prerequisite for predicting the proper orientation of 1-4 in the binding pocket of the tyrosinase enzyme.…”

Hydroxamic Acid as a Potent Metal-Binding Group for Inhibiting Tyrosinase

“…Nevertheless, practically effective strategies for finding new modulators or other targets have been less reported. We have reported novel tyrosinase and laccase inhibitors by examining cross-inhibition between copper-containing enzymes [ 21 ]. Adopting the method of comprehensively cross-examining inhibitors of tyrosinase and other metalloenzymes, including HDACs, may lead to the discovery of tyrosinase inhibitors with new MBGs.…”

“…We have discovered tyrosinase inhibitors using biochemical, biophysical, and computational methods [ 13 , 21 , 22 , 23 , 24 ]. In this study, we report that hydroxamic acid can be a general and strong MBG for inhibiting tyrosinase.…”