Nucleotide Hydrolysis and Electron Transfer Reactions in Nitrogenase Catalysis

Abstract

Nitrogenase is a complex, two component metalloprotein composed of an iron (Fe) protein and a molybdenum-iron (MoFe) protein. The Fe protein is a homodimer (M_r = 64 kDa) that contains two MgATP binding sites and a single [4Fe-4S] cluster bridging the two subunits (Georgiadis et al, 1992). The MoFe protein is an α₂β₂ heterotetramer (M_r = 250 kDa) that contains two pairs of novel metallocenters, called P- or [8Fe-7S] clusters and iron-molybdenum cofactors (FeMoco) (Kim, Rees, 1992). All substrate reduction reactions catalyzed by nitrogenase require the sequential association and dissociation of these two component proteins (Hageman, Burris, 1978), with the concomitant hydrolysis of at least two MgATP molecules coupled to the transfer of a single electron between the proteins (Mortenson et al, 1993). The flow of electrons proceeds from the [4Fe-4S] cluster of the Fe protein to the P-clusters in the MoFe protein (Lanzilotta, Seefeldt, 1996) and finally to FeMoco, (Shah, Brill, 1977), where substrates bind and are reduced. MgATP appears to play a variety of roles in the nitrogenase mechanism which will be discussed in terms of three sequential steps in the reaction mechanism: (i) the binding of MgATP to the Fe protein component of nitrogenase induces protein conformational changes which are a prerequisite for the proper docking of this protein to the MoFe protein, (ii) the hydrolysis of MgATP by the Fe protein-MoFe protein complex is coupled by an unknown mechanism to the transfer of an electron from the Fe protein to the MoFe protein, and finally (iii) the hydrolysis of MgATP to MgADP appears to be involved in triggering dissociation of the Fe protein from the MoFe protein following electron transfer.