Studies on the Mechanism of Electron Bifurcation Catalyzed by Electron Transferring Flavoprotein (Etf) and Butyryl-CoA Dehydrogenase (Bcd) of Acidaminococcus fermentans.

Abstract Electron bifurcation is a fundamental strategy of energy coupling originally discovered in the Q-cycle of many organisms. Recently a flavin-based electron bifurcation has been detected in anaerobes, first in clostridia and later in acetogens and methanogens. It enables anaerobic bacteria and archaea to reduce the two [4Fe-4S] cluster-containing ferredoxin, an energy rich compound that is used to conduct difficult reductions as well as to increase the efficiency of substrate level and electron transport phosphorylations (SLP and ETP). Here we characterize the bifurcating electron transferring flavoprotein (EtfAf) and butyryl-CoA dehydrogenase (BcdAf) from Acidaminococcus fermentans which couple the exergonic reduction of crotonyl-CoA to butyryl-CoA to the endergonic reduction of ferredoxin both with NADH. EtfAf contains one FAD (α-FAD) in subunit α and a second FAD (β-FAD) in subunit β. The distance between the two isoalloxazine rings is 18 Angstrom. The EtfAf-NAD+ complex structure revealed β-FAD as acceptor of the hydride of NADH. The formed β-FADH− is considered as the bifurcating electron donor. Due to a conformational change, α-FAD is able to approach β-FADH− by ca. 5 Angstrom and take up one electron yielding a stable anionic semiquinone, α-FAD●−, which due to a second conformational change donates this electron further to FAD of BcdAf. The remaining non-stabilized neutral semiquinone, β-FADH●, immediately reduces ferredoxin. Repetition of this process affords a second reduced ferredoxin and FADH− of BcdAf that converts crotonyl-CoA to butyryl-CoA.