Coenzyme FO, a deazaflavin chromophore, plays a crucial role as a hydride acceptor/donor in the central methanogenic pathway [1][2].

Coenzyme FO formation is facilitated by dual radical SAM active sites located in either the CofG and CofH enzymes or solely in the FbiC enzyme, which together serve as the operational domains of Fo synthase. In Methanocaldococcus jannaschii, F420 biosynthesis involves eight enzymes, with the production of the deazaflavin chromophore (Fo) being the final unresolved process. Coenzyme F420, a pivotal low-redox-potential electron carrier in methanogenic metabolism, undergoes reduction via hydrogen by F420-dependent hydrogenase. Additionally, it functions as a hydride transfer coenzyme for the F420-specific glucose-6-phosphate dehydrogenase (Fgd) in mycobacteria and is prevalent across all methanogenic and certain non-methanogenic archaea. It plays a crucial role in energy metabolism, NADP reduction, oxygen detoxification, and sulfite reduction. M. tuberculosis utilizes reduced F420 (F420H2) to convert NO2 back to NO, potentially diminishing the antibacterial efficacy of macrophages.