PNNL

Abstract

The enzyme nitrogenase reduces dinitrogen to ammonia utilizing electrons, protons, and energy obtained from the binding and hydrolysis of ATP. Mo-dependent nitrogenase has two symmetric halves each comprising an ATP-dependent reductase, termed the Fe Protein, and a catalytic protein, known as the MoFe-protein, which hosts an electron transfer P-cluster and the active-site metal cofactor (FeMo-co). A series of synchronized events for the electron transfer have been characterized experimentally, in which electron delivery is coupled to nucleotide hydrolysis and regulated by an intricate allosteric network. We report a graph theory analysis of the mechanical coupling in the nitrogenase complex as a key step to understanding the dynamics of allosteric regulation of nitrogen reduction. This analysis shows that regions near the active sites undergo large-scale, large-amplitude correlated motions that are spanned by local motions that enable communications within each half and between the two halves of the complex. Regions in the MoFe protein coupled to the Fe protein were validated against an analysis of the solution phase dynamics of the protein complex via hydrogen-deuterium exchange. These regions include the P-loops and the switch regions in the Fe proteins, the loop containing the residue ?-188Ser adjacent to the P-cluster in the MoFe protein, and the residues near the protein-protein interface. In particular, it is found that: (i) within each Fe protein, the switch region I and II are coupled to the [4Fe-4S] cluster; (ii) within each half of the complex, the switch regions I and II are coupled to the loop containing ß-188Ser; (iii) between the two halves of the complex, the regions near the nucleotide binding pockets of the two Fe proteins (in particular the P-loops, located over 130 Å apart) are also mechanically coupled. Notably, we found that residues next to the P-cluster (in particular the loop containing ?-188Ser) are important for the communication between the two halves.