PURIFICATION AND CHARACTERIZATION OF AN ENZYME THAT CATALYZES RING-CLEAVAGE OF ASPERGILLIC ACID, FROM TRICHODERMA-KONINGII ATCC-76666

The aspergillic acid degrading enzyme (ADE) that catalyzes the cleavag e of the pyrazine ring in aspergillic acid (AA, 1-hydroxy-3-isobutyl-6 -sec-butyl-2-pyrazinone) was purified to electrophoretic homogeneity f rom extracts of Trichoderma koningii ATCC 76666. ADE was a homodimeric protein with a molecular mass of 112 kDa, contained 1 mol of FAD per mol of subunit, and required NAD(P)H and molecular oxygen for its acti vity. ADE had an isoelectric point of around 5.3, and an optimum pH of 7.0-8.0. p-Chloromercuribenzoate and HgCl2 completely inhibited ADE a ctivity, while metal chelating reagents, alpha,alpha'-dipyridyl and o- phenanthroline, were not inhibitors. The substrate specificity among A A-related compounds was that hydroxyaspergillic acid was a poor substr ate (16% of the activity for AA) and deoxyaspergillic acid did not ser ve as a substrate.