Mössbauer evidence for exchange-coupled siroheme and [4Fe-4S] prosthetic groups in Escherichia coli sulfite reductase. Studies of the reduced states and of a nitrite turnover complex.

We have studied the hemoprotein subunit (SiR) of Escherichia coli NADPH-sulfite reductase with Mössbauer spectroscopy in a variety of complexes and oxidation states. We demonstrated earlier for oxidized SiR (Christner, J. A., Münck, E., Janick, P. A., and Siegel, L. M. (1981) J. Biol. Chem. 256, 2098-2101) that the two metal centers, a siroheme and a [4Fe-4S] cluster, are covalently linked as witnessed by the observation of strong exchange-coupling. The studies reported here show that the centers remain coupled in the one- and two-electron-reduced states of SiR. Exchange-coupling is also observed for the turnover complex, SiR X NO; this species can be prepared by reacting fully reduced SiR, which can function as a nitrite reductase, with nitrite. The coupling is also maintained in the presence of certain chaotropic agents. In SiR X NO, the exchange interactions between the S = 1/2 Fe(II) X siroheme X NO and the iron-sulfur cluster induce paramagnetism at the iron sites of the [4Fe-4S] cluster which is formally in the (diamagnetic) 2+ oxidation state. Analysis of the Mössbauer spectra shows that the iron sites of the cluster are equivalent in pairs, which are distinguishable by positive and negative magnetic hyperfine coupling constants. This study has revealed that the Mössbauer parameters of the siroheme, an Fe X isobacteriochlorin, are very similar to those observed for other hemes. The data obtained for the reduced forms of uncomplexed enzyme show that the siroheme iron is Fe(II), that it is paramagnetic, and possibly in the S = 1 intermediate spin state.