Radical S-Adenosyl-l-Methionine (SAM) Enzyme Involved in the Maturation of the Nitrogenase Cluster.

Nitrogenase is the only known enzymatic system that converts atmospheric dinitrogen (N) into bioavailable ammonia (NH). The active-site cofactor responsible for this reactivity is a [(R-homocitrate)MoFeSC] cluster that is designated as the M-cluster. This important cofactor is assembled stepwise from a pair of [FeS] clusters that become fused into a [FeSC] core before additional refinements take place to complete the biosynthesis.

Formation of a homocitrate-free iron-molybdenum cluster on NifEN: implications for the role of homocitrate in nitrogenase assembly.

Molybdenum (Mo)-dependent nitrogenase is a complex metalloprotein that catalyzes the biological reduction of dinitrogen (N(2)) to ammonia (NH(3)) at the molybdenum-iron cofactor (FeMoco) site of its molybdenum-iron (MoFe) protein component. Here we report the formation of a homocitrate-free, iron-molybdenum ("FeMo") cluster on the biosynthetic scaffold of FeMoco, NifEN. Such a NifEN-associated "FeMo" cluster exhibits EPR features similar to those of the NifEN-associated, fully-complemented "FeMoco", which originate from the presence of Mo in both cluster species; however, "FeMo" cluster and "FeMoco" display different temperature-dependent changes in the line shape and the signal intensity of their respective EPR features, which reflect the impact of homocitrate on the redox properties of these clusters.

Insertion of heterometals into the NifEN-associated iron-molybdenum cofactor precursor.

The cofactors of Mo-, V-, Fe-dependent nitrogenases are believed to be highly homologous in structure despite the different types of heterometals (Mo, V, and Fe) they contain. Previously, a precursor form of the FeMo cofactor (FeMoco) was captured on NifEN, a scaffold protein for FeMoco biosynthesis. This all-Fe precursor closely resembles the Fe/S core structure of the FeMoco and, therefore, could reasonably serve as a precursor for all nitrogenase cofactors.