New Electron-Transfer Chain to a Flavodiiron Protein in <i>Fusobacterium nucleatum</i> Couples Butyryl-CoA Oxidation to O₂ Reduction

<i>Fusobacterium nucleatum</i>, a Gram-negative obligate anaerobe, is common to the oral microbiota, but the species is known to infect other sites of the body where it is associated with a range of pathologies. At present, little is known about the mechanisms by which <i>F. nucleatum</i> mitigates against oxidative and nitrosative stress. Inspection of the <i>F. nucleatum</i> subsp. <i>polymorphum</i> ATCC 10953 genome reveals that it encodes a flavodiiron protein (FDP; FNP2073) that is known in other organisms to reduce NO to N₂O and/or O₂ to H₂O. FNP2073 is dicistronic with a gene encoding a multicomponent enzyme termed BCR for <u>b</u>utyryl-<u>C</u>oA <u>r</u>eductase. BCR is composed of a butyryl-CoA dehydrogenase domain (BCD), the C-terminal domain of the α-subunit of the electron-transfer flavoprotein (Etfα), and a rubredoxin domain. We show that BCR and the FDP form an α₄β₄ heterotetramic complex and use butyryl-CoA to selectively reduce O₂ to H₂O. The FAD associated with the Etfα domain (α-FAD) forms red anionic semiquinone (FAD^•-), whereas the FAD present in the BCD domain (δ-FAD) forms the blue-neutral semiquinone (FADH^•), indicating that both cofactors participate in one-electron transfers. This was confirmed in stopped-flow studies where the reduction of oxidized BCR with an excess of butyryl-CoA resulted in rapid (<1.6 ms) interflavin electron transfer evidenced by the formation of the FAD^•-. Analysis of bacterial genomes revealed that the dicistron is present in obligate anaerobic gut bacteria considered to be beneficial by virtue of their ability to produce butyrate. Thus, BCR-FDP may help to maintain anaerobiosis in the colon.