Structure-function studies of a laccase from Melanocarpus albomyces

Laccases are copper metalloenzymes, which catalyze the oxidation of a variety of aromatic compounds. The one-electron oxidation of the reducing substrate occurs concomitantly with a four-electron reduction of molecular oxygen to water. As laccases do not need an added coenzyme but use molecular oxygen, they are a very potent enzyme group for large-scale industrial use. Laccases have been tested in pulp bleaching, textile dye decolorization, bioglueing and detoxification. We have earlier determined the three-dimensional structure of Melanocarpus albomyces laccase (MaL) at 2.4 Angstrom resolution. MaL consists of 559 amino acids and four copper atoms. The three-dimensional structure of MaL revealed an overall fold similar to other known laccase structures. An enzyme consists of three similar domains with an anti-parallel -sheet folded into a barrel-like structure. Copper atoms form the mono- (one type-1 Cu) and trinuclear sites (one type-2 Cu and two type-3 Cu). Aromatic substrates are oxidized near the mononuclear site and the electrons are transferred to the trinuclear site, where a molecular oxygen is reduced to water molecules. Despite the fact that the overall fold of MaL was similar to other known laccases, the major differences were observed in the loops forming the substrate-binding site for the aromatic compounds. Mal also had a dissimilar C-terminus blocking the access to the trinuclear site. Other known laccases have a wide cleft, suggesting that oxygen molecules enter and water molecules exit through this cleft. The C-terminus of MaL formed a plug in the putative oxygen tunnel. In addition, the binding of dioxygen to the trinuclear site of MaL was observed. We have recently collected a new native data set at 1.8 Å resolution at the Hamburg synchrotron radiation source. The new high resolution structure reveal new details about both ligand binding sites of MaL