EVIDENCE FOR AN IRON CENTER IN THE AMMONIA MONOOXYGENASE FROM NITROSOMONAS-EUROPAEA

Binding of the ligand, nitric oxide, in the presence of reductant was used to identify a ferrous S = 3/2 signal, characteristic of a ferrous nitrosyl complex, and a g = 2.03 copper or iron signal in membranes o f the ammonia-oxidizing bacterium, Nitrosomonas europaea. The same fer rous S = 3/2 signal is thought to be a component of the membrane-assoc iated methane monooxygenase (pMMO) of Methylococcus capsulatus Bath, s ince it is seen in the membrane fraction of cells expressing pMMO and in the purified enzyme, but not in the membrane fraction of cells expr essing the soluble MMO [Zahn, J.A. and DiSpirito, A.A. (1996) J. Bacte riol. 178, 1018-1029]. Treatment of resting membranes or cells of N. e uropaea with nitrapyrin, 2-chloro,6-trichloromethylpyridine, resulted in the increase in magnitude of a g = 6, high-spin ferric iron signal. In the presence of NO and reductant, nitrapyrin prevented the formati on of the S = 3/2 nitrosyl-iron complex while increasing the intensity of the g = 6 signal. Nitrapyrin is a specific inhibitor of, and is re duced by, the ammonia monoxygenase (AMO) [Bedard, C. and Knowles, R. ( 1989) Microbiol. Rev. 53, 68-83]. Taken together the data suggest that iron capable of forming the S = 3/2 complex is a catalytic component of AMO of N. europaea, possibly a part of the oxygen-activating center . Inactivation of the membrane-associated AMO with acetylene did not d iminish the S = 3/2 nitrosyl-iron signal, the g = 6 signal, or the g = 6 signal.