PURIFICATION AND PROPERTIES OF EMODIN DEOXYGENASE FROM PYRENOCHAETA-TERRESTRIS

Emodin deoxygenase, which catalyses the reduction of emodin to chrysophanol, was purified 17-fold from crude extracts of Pyrenochaeta terrestris. The Mr of the enzyme was 103 000. Upward curvature was exhibited by the plot of rate vs concentration of the crude extract. The protein fraction obtained from gel filtration with Sephadex G-75 was activated by ATP plus a low Mr fraction; ATP alone or low Mr fraction alone did not increase its activity. The Km for NADPH of the crude extract was 3 μM, that after NADPH gel filtration of the crude extract was 1.5 mM. It is proposed that ATP plus an unidentified factor increase emodin deoxygenase activity by lowering the Km for NADPH. Iron II, which increased activity of the crude extract, inhibited activity of the partially purified enzyme by 95%. Sulphydryl reagents inhibited activity by 90%. Partially purified emodin deoxygenase activity was low in the absence of mercaptans and was increased eight-fold by the addition of dithiothreitol. It is proposed that a pair of thiol groups is required for activity and that they occur in the disulphide form in the absence of mercaptans. © 1990.