An all-ferrous state of the Fe protein of nitrogenase. Interaction with nucleotides and electron transfer to the MoFe protein.

The MoFe protein of nitrogenase catalyzes the six-electron reduction of dinitrogen to ammonia. It has long been believed that this protein receives the multiple electrons it requires one at a time, from the [4Fe-4S]2+/+ couple of the Fe protein. Recently an all-ferrous [4Fe-4S]0 state of the Fe protein was demonstrated suggesting instead a series of two electron steps involving the [4Fe-4S]2+/0 couple. We have examined the interactions of the [4Fe-4S]0 Fe protein with nucleotides and its ability to transfer electrons to the MoFe protein. The [4Fe-4S]0 Fe protein binds both MgATP and MgADP and undergoes the MgATP induced conformational change and then binds properly to the MoFe protein, as evidenced by the fact that the behavior of the 0 and +1 oxidation states in the chelation and chelation protection assays are indistinguishable. Nucleotide binding does not effect the distinctive UV/Vis, CD, or Mössbauer spectra exhibited by the [4Fe-4S]0 Fe protein; however, because the intensity of the g = 16.4 EPR signal of the [4Fe-4S]0 Fe protein is extremely sensitive to minor variations of the rhombicity parameter E/D, the EPR signal is sensitive to the binding of nucleotides. A 50:50 mixture of [4Fe-4S]2+ and [4Fe-4S]0 Fe protein results in electron self-exchange and 100% production of [4Fe-4S]+ Fe protein, demonstrating that the +1/0 couple is fully reversible. MgATP is absolutely required for electron transfer from the [4Fe-4S]0 Fe protein to the reduced state of the MoFe protein. In that reaction both electrons are transferred and are used to reduce substrate.