The inactivation of dihydroxy-acid dehydratase in Escherichia coli treated with hyperbaric oxygen occurs because of the destruction of its Fe-S cluster, but the enzyme remains in the cell in a form that can be reactivated.
The enzyme dihydroxy-acid dehydratase previously has been shown to be inactivated in vivo in Escherichia coli within minutes of exposure to hyperbaric O2. In this paper, we show its inactivation is due to the destruction of its catalytically active [4Fe-4S] cluster. The inactivation is not followed by an appreciable decrease in the amount of dihydroxy-acid dehydratase protein as determined by Western blots. Thus, the protein from the inactivated enzyme remains unproteolyzed in the cells. Dihydroxy-acid dehydratase activity recovers after the cells treated with hyperbaric O2 are returned to ambient oxygen. Since this recovery in activity is not accompanied by a significant increase in dihydroxy-acid dehydratase protein and is not prevented by chloramphenicol, it appears primarily to be due to reactivation of the previously inactivated enzyme. The reactivation occurs by reconstitution of the enzyme's Fe-S cluster. These results demonstrate that this enzyme can cycle between forms in which the Fe-S cluster is either present or absent. The facile ability to cycle between these two forms would be compatible with a regulatory role in addition to a catalytic role for this enzyme.
Flint
D H
DH
Central Research and Development Department, E. I. du Pont de Nemours and Company, Wilmington, Delaware 19880.
Smyk-Randall
E
E
Tuminello
J F
JF
Draczynska-Lusiak
B
B
Brown
O R
OR
D002701:Chloramphenicol / Q000633:toxicity; D002852:Chromatography, Ion Exchange; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000254:growth & development; D006836:Hydro-Lyases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D006931:Hyperbaric Oxygenation; D007501:Iron / Q000032:analysis; D007506:Iron-Sulfur Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D010100:Oxygen / Q000494:pharmacology*; D011312:Pressure
D007506:Iron-Sulfur Proteins; D002701:Chloramphenicol; D007501:Iron; D006836:Hydro-Lyases; C020052:dihydroxyacid dehydratase; D010100:Oxygen