Inactivation of Tyrosinase Photoinduced by Folic Acid and its Photoproducts

ZURBANO,BEATRIZ N.; DANTOLA, MARIA LAURA; THOMAS, ANDRÉS H.

Córdoba

Congreso; 16th International Congress on Photobiology; 2014

International Union of Photobiology

UV-A radiation (320-400 nm) can induce damage to biomacromolecules through photosensitized reactions. This indirect action may be mediated by endogenous or exogenous photosensitizers and can take place via different mechanisms [1].  UV-A excitation of folic acid (PteGlu), a conjugated pterin, leads to its oxidation, yielding 6-formylpterin (Fop) and p-aminobenzoylglutamic acid as photoproducts. Hydrogen peroxide (H2O2) is formed during this reaction. In turn, Fop is transformed into 6-carboxypterin (Cap) upon further photooxidation.  Vitiligo is a skin disease that causes lack of pigmentation due to inactivation of enzymes of the biosynthesis of melanin. In the skin of these patients there is an accumulation of pterin derivatives with high levels of H2O2 [2]. Tyrosinase in mammals catalyzes the first and rate-limiting step in melanogenesis. Recent studies suggested that pterin (Ptr), the parent and unsubstituted compound of oxidized pterins, photoinduces the inactivation of tyrosinase [3].  In this work, we investigate the damage of tyrosinase photoinduced by PteGlu and its photoproducts. After UV-A excitation, aqueous solutions containing the enzyme and the photosensitizer (PteGlu, Fop, Cap, Ptr) were analyzed by UV-Vis spectrophotometry, enzyme activity measurement, fluorescence spectroscopy and High-performance Liquid Chromatography.  The results indicated that PteGlu and its photoproducts can fotoinactivate the tyrosinase. The most efficient photosensitizer was Fop. The photoinactivation takes place through two differente pathways: i) a photochemical process initiated by an electron-transfer from the enzyme to the triplet excited states of pterins and ii) the oxydation of the enzyme by the H2O2 produced during the photooxidation of PteGlu and its photoproduct. The photochemical process would lead to an unspecific oxydation of the protein, whereas the reaction with H2O2 would act only on the active site.  Acknowledgments. Authors thank CONICET, ANPCyT and UNLP for financial support.  [1] J.Cadet , et. al, Mutat. Res., 2005, 3, 571. [2] K. U. Schallreuter, et. al., Science, 1994, 263, 1444. [3] M. L.Dántola, et. al., Biochem. Biophys. Res. Commun.,2012, 424, 568.