Human phenylalanine hydroxylase is activated by H2O2:: a novel mechanism for increasing the L-tyrosine supply for melanogenesis in melanocytes

Epidermal phenylalanine hydroxylase (PAH) produces L-tyrosine from the essential amino acid L-phenylalanine supporting melanogenesis in human melanocytes. Those PAH activities increase linearly in the different skin phototypes I-VI (Fitzpatrick classification) and also increase up to 24 h after UVB light with only one minimal erythemal dose. Since UVB generates also H2O2, We here asked the question whether this reactive oxygen species could influence the activity of pure recombinant human PAH. Under saturating conditions with the substrate L-phenylalanine (1 x 10(-3) M), the V-max for enzyme activity increased 4-fold by H2O2 (>2.0 x 10(-3) M). Lineweaver-Burk analysis identified a mixed activation mechanism involving both the regulatory and catalytic domains of PAR Hyperchem molecular modelling and Deep View analysis support oxidation of the single Trp(120) residue to 5-OH-Trp(120) by H2O2 causing a conformational change in the regulatory domain. PAH was still activated by H2O2 in the presence of the electron donor/cofactor 6(R)-L-erythro-5,6,7,8-tetrahydrobiopterin despite slow oxidation of this cofactor. In vivo FT-Raman spectroscopy confirmed decreased epidermal phenylalanine in association with increased tyrosine after UVB exposure. Hence, generation of H2O2 by UVB can activate epidermal PAH leading to an increased L-tyrosine pool for melanogenesis. (C) 2004 Elsevier Inc. All rights reserved.