Photoinactivation of tyrosinase sensitized by folic acid photoproducts.

Tyrosinase catalyzes in mammals the first and rate-limiting step in the biosynthesis of the melanin, the main pigment of the skin. Pterins, heterocyclic compounds able to photoinduce oxidation of biomolecules, accumulate in the skin of patients suffering from vitiligo, where there is a lack of melanin. Folic acid (PteGlu) is a conjugated pterin widespread in biological systems. Aqueous solutions of tyrosinase were exposed to UV-A irradiation (350nm) in the presence of PteGlu and its photoproducts (6-formylpterin and 6-carboxypterin). The reactions were followed by UV-Vis spectrophotometry, enzyme activity measurement, fluorescence spectroscopy and HPLC. In this work, we present data that demonstrate unequivocally that solutions of tyrosinase exposed to UV-A irradiation in the presence of PteGlu, undergo enzyme inactivation. However, PteGlu itself causes a negligible effect on the activity of the enzyme. In contrast, PteGlu photoproducts are efficient photosensitizers. The tyrosinase inactivation involves two different pathways: (i) a photosensitization process and (ii) the oxidation of the enzyme by the hydrogen peroxide produced during the photooxidation of PteGlu and its photoproduct. The former pathway affects both the active site and the tryptophan residues, whereas the latter affects only the active site. The biological implications of the results are discussed.
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