Interactions that shape the complementary reactivities of the flavins of bifurcating electron transfer flavoproteins

Flavin‐based electron bifurcation accomplishes the feat of producing highly‐reducing electrons at the expense of a less reactive source. By consuming two modestly‐reducing electrons from NADH for each one electron transferred to the more reducing ferredoxin acceptor, the process conforms to the laws of thermodynamics. Critical mechanistic ingredients include coupling between two separate electron transfer reactions, a protein conformation change preventing the high‐energy electron from exploiting the more exergonic path, and high‐efficiency transfer of that electron to the more reducing acceptor instead. All of this is accomplished based on the dynamic structure of the protein, in conjunction with the distinctive electrochemical reactivity of flavins. We are combining QM/MM approaches with site‐directed mutagenesis and electrochemistry to clarify how the protein enforces one‐electron chemistry at the electron transfer site but bifurcation at the other. Complementary studies seek to identify covalent modifications that accumulate on one of the two flavins, especially at high pH.