Mössbauer Studies on the Active Fe …[2Fe‐2S] Site of Putidamonooxin, Its Electron Transport and Dioxygen Activation Mechanism

Abstract

Putidamonooxin, the oxygenase of a 4‐methoxybenzoate monooxygenase enzyme system, catalyzes the oxidative O‐demethylation of the substrate 4‐methoxybenzoate in conjunction with the NADH: putidamonooxin oxidoreductase. Putidamonooxin is a conjugated iron‐sulfur protein which needs iron ions as cofactors for its enzymatic activity. Putiamonooxin was isolated from Pseudornonus putida, which was grown on a 57Fe‐enriched culture medium. Thus putidamonooxin was enriched in vivo with 57Fe up to about 80%. During our Mössbauer study of putidamonooxin a number of parameters have been varied: (a) the oxidation state of putidamonooxin (oxidized, reduced and aerobically reoxidized); (b) the substrate bound to putidamonooxin (Cmethoxybenzoate, benzoate, 4‐tert‐butylbenzoate); (c) the temperature between 2.7 K and 245 K; (d) the applied magnetic field between 0 and 0.1 T and (e) the amount of iron cofactor. From our Mössbauer results it is obvious that the iron‐sulfur centers of putidamonooxin are [2 Fe‐2 S] clusters similar to those of the plant‐type ferredoxins. Further, we have evidence for the existence of iron ions (one per [2 Fe‐2S] cluster), which serve as cofactors for the dioxygen activation, functioning as the dioxygen binding site and mediating the electron flow from the [2 Fe‐2 S] cluster to dioxygen. Copyright © 1981, Wiley Blackwell. All rights reserved