Microorganisms utilize electron bifurcating enzymes to carry out thermodynamically unfavorable
reactions where a single enzyme reversibly couples an endergonic reaction to an exergonic
reaction to generate a net reversible reaction with minimal free energy change. In essence, an
exergonic chemical reaction is used to drive an endergonic one. Electron bifurcation is now
recognized as a major energy coupling system in biology although only a limited number of
examples are known. We very recently discovered a large and diverse family of bifurcating
enzymes, termed Bfu, that is surprisingly ubiquitous in the microbial world. Although very few
Bfu enzymes have been characterized, they are predicted to catalyze reactions involving an
extensive range of substrates not previously known to be involved in electron bifurcation
reactions. Our overriding hypotheses are that 1) Bfu enzymes employ a new bifurcation
mechanism that uses a combination of flavin and iron sulfur clusters, 2) the mechanism and
associated electron transfer pathways allow tremendous flexibility in terms of the range of
substrates that can be used by Bfu members, and 3) numerous types of Bfu enzyme exist that
have yet to be characterized, including membrane-bound ion-translocating complexes, and some
others appear to couple electron bifurcation to non-redox cellular processes.
The specific aims of the proposed research are 1) to determine the unique properties of the
bifurcating site in Bfu enzymes and elucidate the mechanism of electron bifurcation, 2) to
characterize new and unprecedented types of bifurcating Bfu enzymes including those that
potentially utilize substrates such as hydrogen peroxide and acetylene, and ones involved with
cellular processes, such as proteolysis, and 3) to characterize a membrane-bound iontranslocating
Bfu family member that is proposed to couple electron bifurcation to the formation
of chemical gradients. Our objectives will be achieved by utilizing fermentations of
hyperthermophilic and thermophilic microbes, recombinant production and anaerobic purification
of cytoplasmic and membrane enzyme complexes, various biochemical and kinetic techniques,
and UV/visible and EPR spectroscopy. This project will also leverage an on-going collaboration
using cryo-EM to determine structures of electron bifurcating enzymes.
Interested? Email Mike Adams (adamsm@uga.edu), University of Georgia