The three proteins that play a direct role in the M-cluster synthesis are NifH, NifEN, and NifB. The NifB protein is responsible for the assembly of the Fe-S core of the cofactor; a process that involves stitching together two [4Fe-4S] clusters. NifB belongs to the SAM (S-adenosyl-L-methionine) enzyme superfamily. During the biosynthesis of the FeMo cofactor, NifB and its SAM cofactor are directly involved in the insertion of a carbon atom at the center of the Fe-S complex. An equivalent of SAM donates a methyl group, which becomes the interstitial carbide of the M-cluster. The methyl group of SAM is mobilized by radical removal of an H by a 5’-deoxyadenosine radical (5’-dA·). Presumably, a transient –CH2· radical is formed that is subsequently incorporated into the metal cluster forming a Fe-carbide species. The interstitial carbon remains associated with the FeMo cofactor after insertion into the nitrogenase, The nature of the central atom in FeMoco as a carbon species was identified in 2011. The approach for the identification relied on a combination of C/N-labeling and pulsed EPR spectroscopy as well as X-ray christallographic studies at full atomic resolution. Additionally, X-ray diffractometry was used to verify that there was a central carbon atom in the middle of the FeMo cofactor and x-ray emission spectroscopic studies showed that central atom was carbon due to the 2p→1s carbon-iron transition. The use of X-ray crystallography showed that while the FeMo cofactor is not in its catalytic form, the carbon keeps the structure rigid which helps describe the reactivity of nitrogenase.