Nitrogenase XII. Mössbauer studies of the MoFe protein from Clostridium pasteurianum W5

We have studied the molybdenum-protein (MoFe protein) from Clostridium pasteurianum with Mössbauer spectroscopy in the temperature range from 1.5 to 200 K in magnetic fields up to 55 kG. Except for some small differences in the hyperfine parameters the results for the C. pasteurianum protein are essentially the same as those published previously for the protein from Azotobacter vinelandii, i.e. (30 +/- 2) Fe atoms partition into two identical cofactor centers M (each center most likely containing six Fe atoms and one Mo atom), four P-clusters (each center containing four Fe atoms), and one iron environment labeled S (about two Fe atoms per holoenzyme). We have analyzed the spectra of the cofactor centers in three distinct oxidation states, Formula: (see test). The diamagnetic (electronic spin S = 0) state MOX is attained by oxidation of the native, EPR-active (S = 3/2) state MN. The reduced state MR is observed in steady state under nitrogen fixing conditions; high-field Mössbauer studies show that the cofactor centers are paramagnetic (integer electronic spin S greater than or equal to 1) in the state MR. We have evaluated the complex high-field spectra resulting from the P-clusters in the oxidized state POX. The analysis shows that one iron site is characterized by a positive hyperfine coupling constant A0 while the other three sites have A0 less than 0. A slightly modified set of parameters also fits the high-field data of the MoFe protein from A. vinelandii. Finally, we will present a discussion summarizing our principle results obtained to date for the proteins from A. vinelandii and C. pasteurianum.