Characterization of the [omega]-hydroxylase of Pseudomonas oleovorans as a nonheme iron protein

Abstract

The [omega]-hydroxylase of Pseudomonas oleovorans, which catalyzes the hydroxylation of fatty acids and alkanes and the epoxidation of alkenes in the presence of a reduced pyridine nucleotide, a reductase, rubredoxin, and molecular oxygen, has been purified to electrophoretic homogeneity. Octane hydroxylation and octadiene epoxidation activities appear to remain at a constant ratio during the purification procedure. The hydroxylase has been characterized as a nonheme iron protein containing one iron atom and one cysteine residue per polypeptide chain of molecular weight 40,800. The enzyme is inhibited by cyanide, and activity is restored upon removal of the cyanide by dialysis. Iron is removed from the enzyme by dialysis against EDTA provided that a reducing agent such as dithionite or ascorbate is also added, and enzyme activity is restored by the addition of ferrous ions to the apohydroxylase.