Ferrisiderophore reductase activity in Bacillus megaterium

Abstract

The release of Fe from ferrisiderophores (microbial ferric-chelating Fe transport cofactors) by cell-free extracts of B. megaterium was demonstrated. Reductive transfer of iron from ferrisiderophores to the ferrous-chelating agent ferrozine was measured spectrophotometrically. This ferrisiderophore reductase activity (NADPH:ferrisiderophore oxidoreductase) was associated primarily with the cell soluble rather than particulate (membrane) fraction. Ferrisiderophore reductase was inhibited by O2 and required the addition of a reductant (NADPH was most effective) for maximal activity. The activity was destroyed by both heat and protease treatments and was inhibited by iodoacetamide treatment. Ferrisiderophore reductase activity for several microbial ferrisiderophores was measured; highest activity was displayed for ferrischizokinen, the ferrisiderophore produced by this organism. The Km and Vmax values of the reductase for ferrischizokinen were 2.5 .times. 10-4 M and 35.7 nmol/min per mg of protein, respectively. The addition of deferrisiderophore reduced the velocity of the ferrisiderophore reductase reaction. Preliminary fractionation of the cell-soluble material by gel filtration chromatography resulted in the demonstration of ferrisiderophore reductase activity in 3 peaks of different MW. Ferrisiderophore reductase probably mediates entrance of Fe into cellular metabolism.