EPR of hydroxylamine oxidoreductase from Nitrosomonas europaea

Abstract

EPR spectra are reported for the multiheme enzyme hydroxylamine oxidoreductase of Nitrosomonas europaea. Signals arising from several different types of low-spin ($\text{S =}\text{1}\text{2}$) ferric heme were observed in the resting (oxidized) enzyme, but no evidence was obtained for high-spin ($\text{S =}\text{5}\text{2}$) heme, copper or iron-sulfur centers. While overlap of the low-spin heme signals at X-band frequency complicates complete assignment of $\text{g}$ values, four species with low field peaks at $\text{g}$ 3.4, 3.1, 3.0 and 2.7 were clearly resolved. Complete reduction of the enzyme with sodium dithionite abolished all major signals. Partial reduction of the enzyme by hydroxylamine caused disappearance of signals at $\text{g 3.1, g 2.22}$ and $\text{g 1.35}$ indicating selective reduction of this low-spin component by the substrate.