MgATP-bound and nucleotide-free structures of a nitrogenase protein complex between the Leu 127Δ-Fe-protein and the MoFe-protein

A mutant form of the nitrogenase iron protein with a deletion of residue Leu 127, located in the switch II region of the nucleotide binding site, forms a tight, inactive complex with the nitrogenase molybdenum iron (MoFe) protein in the absence of nucleotide. The structure of this complex generated with proteins from Azotobacter vinelandii (designated the L127 Delta -Av2-Av1 complex) has been crystallographically determined in the absence of nucleotide at 2.2 Angstrom resolution and with bound MgATP (introduced by soaking) at 3.0 Angstrom resolution. As observed in the structure of the complex between the wild-type A. vinelandii nitrogenase proteins stabilized with ADP . AlF4-, the most significant conformational changes in the L127 Delta complex occur in the Fe-protein component. While the interactions at the interface between the MoFe-protein and Fe-proteins are conserved in the two complexes, significant differences are evident at the subunit-subunit interface of the dimeric Fe-proteins, with the L127 Delta -Av2 structure having a more open conformation than the wild-type Av2 in the complex stabilized by ADP . AIF(4)(-). Addition of MgATP to the L127 Delta -Av2-Av1 complex results in a further increase in the separation between Fe-protein subunits so that the structure more closely resembles that of the wild-type, nucleotide-free, uncomplexed Fe-protein, rather than the Fe-protein conformation in the ADP . AIF(4)(-) complex. The L127 Delta mutation precludes key interactions between the Fe-protein and nucleotide, especially, but not exclusively, in the region corresponding to the switch II region of G-proteins, where the deletion constrains Gly 128 and Asp 129 from forming hydrogen bonds to the gamma -phosphate and activating water for attack on this group, respectively. These alterations account for the inability of this mutant to support mechanistically productive ATP hydrolysis. The ability of the L127 Delta -Av2-Av1 complex to bind MgATP demonstrates that dissociation of the nitrogenase complex is not required for nucleotide binding.