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Tyrosinase catalyzes the conversion of phenolic compounds into their quinone derivatives, which are precursors for the formation of melanin, a ubiquitous pigment in living organisms. Because of its importance for browning reactions in the food industry, the tyrosinase from the mushroom Agaricus bisporus has been investigated in depth. In previous studies the tyrosinase enzyme complex was shown to be a H(2)L(2) tetramer, but no clues were obtained of the identities of the subunits, their mode of association, and the 3D structure of the complex. Here we unravel this tetramer at the molecular level. Its 2.3 Å resolution crystal structure is the first structure of the full fungal tyrosinase complex. The complex comprises two H subunits of ∼392 residues and two L subunits of ∼150 residues. The H subunit originates from the ppo3 gene and has a fold similar to other tyrosinases, but it is ∼100 residues larger. The L subunit appeared to be the product of orf239342 and has a lectin-like fold. The H subunit contains a binuclear copper-binding site in the deoxy-state, in which three histidine residues coordinate each copper ion. The side chains of these histidines have their orientation fixed by hydrogen bonds or, in the case of His85, by a thioether bridge with the side chain of Cys83. The specific tyrosinase inhibitor tropolone forms a pre-Michaelis complex with the enzyme. It binds near the binuclear copper site without directly coordinating the copper ions. The function of the ORF239342 subunits is not known. Carbohydrate binding sites identified in other lectins are not conserved in ORF239342, and the subunits are over 25 Å away from the active site, making a role in activity unlikely. The structures explain how calcium ions stabilize the tetrameric state of the enzyme.

The total free radical scavenger capacity (RSC) of 57 edible oils from different sources was studied: olive (24 brands of oils), sunflower (6), safflower (2), rapeseed (3), soybean (3), linseed (2), corn (3), hazelnut (2), walnut (2), sesame (2), almond (2), mixture of oils for salad (2), "dietetic" oil (2), and peanut (2). Olive oils were also studied according to their geographical origins (France, Greece, Italy, Morocco, Spain, and Turkey). RSC was determined spectrophotometrically by measuring the disappearance of the radical 2,2-diphenyl-1-picrylhydrazyl radical (DPPH(*)) at 515 nm. The disappearance of the radical followed a double-exponential equation in the presence of oils and oil fractions, which suggested the presence of two (fast and slow) groups of antioxidants. RSC was studied for the methanol-soluble phase ("methanolic fraction", MF) of the oil, the fraction nonsoluble in methanol ("lipidic fraction", LF), and the nonfractionated oil ("total oil"; TF = MF + LF). Only olive, linseed, rapeseed, safflower, sesame, and walnut oils showed significant RSC in the MF due to the presence of phenolic compounds. No significant differences were found in the RSC of olive oils from different geographical origins. Upon heating at 180 degrees C the apparent constant for the disappearance of RSC (k(T)) and the half-life (t1/2) of RSC for MF, LF, and TF were calculated. The second-order rate constants (k2) for the antiradical activity of some phenolic compounds present in oils are also reported.

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