NifI inhibits nitrogenase by competing with Fe protein for binding to the MoFe protein

Reduction of substrate by nitrogenase requires direct electron transfer from the Fe protein to the MoFe protein. Inhibition of nitrogenase activity in Methanococcus maripaludis occurs when the regulatory protein NifI_1,2 binds the MoFe protein. This inhibition is relieved by 2-oxoglutarate. Here we present evidence that NifI_1,2 binding prevents association of the two nitrogenase components. Increasing amounts of Fe protein competed with NifI_1,2, decreasing its inhibitory effect. NifI_1,2 prevented the co-purification of MoFe protein with a mutant form of the Fe protein that forms a stable complex with the MoFe protein, and NifI_1,2 was unable to bind to an -stabilized Fe protein:MoFe protein complex. NifI_1,2 inhibited ATP- and MoFe protein-dependent oxidation of the Fe protein, and 2OG relieved this inhibition. These results support a model where NifI_1,2 competes with the Fe protein for binding to MoFe protein and prevents electron transfer.