PURIFICATION AND CHARACTERIZATION OF EMODINANTHRONE OXYGENASE FROM ASPERGILLUS-TERREUS

Emodinanthrone oxygenase catalyses the fixation of molecular oxygen into emodinanthrone to give the anthraquinone emodin. The reaction proceeds without external electron donors. The enzyme activity was first detected in crude extracts of the fungus Aspergillus terreus IMI 16043 which produces the seco-anthraquinone, (+)-geodin. Emodinanthrone oxygenase from A. terreus was found to be membrane bound, could be solubilized by non-ionic detergents, and was activated by phospholipids. Although the instability and heterogeneity of the solubilized enzyme retarded its purification, emodinanthrone oxygenase was purified by chromatography on DEAE-cellulose, Hydroxyapatite, Q Sepharose, HPLC gel filtration, and Mono Q columns. The purified enzyme showed a single band with a M(r) of 52000 on SDS-PAGE. The fact that the enzyme was inhibited by ferrous chelating reagents like o-phenanthroline in the presence of reducing reagents suggested that non-haem ferric iron is the catalytic cofactor of the enzyme. Based on these properties. a mechanism for the emodinanthrone oxygenase reaction is proposed.