Speaker
Description
Electron transfer bifurcation ('bifurcation') enables enzymes to produce a more potent reductant at the expense of a weaker one, in apparent defiance of thermodynamics. However no rules are broken, because only one super-reducing electron is produced from each pair of electrons consumed. In-effect, exergonic transfer of the second electron pays for endergonic transfer of the first. Thus, along with the valuable increased-potency reduced electron carrier, an energy-depleted product is also formed. The potent reduced ferredoxin or flavodoxin product supports biological fixation of Nsub2/sub or COsub2/sub, two reactions we would like to be able to facilitate in man-made materials. Hence our interest in understanding the mechanism of bifurcation. A central problem is to understand how bifurcating enzymes provide an exergonic path to allow the favorable electron transfer required to 'pay the bills', but limit its use to a single electron of each pair. Somehow, one electron can (and must) use this path, but the other one is prevented from doing so, being transferred instead to a less favorable acceptor. Successful direction of this electron (and most of the energy) to the unfavorable acceptor is the process that captures energy released from the favorable transfer, making it available for future use.
Working with bifurcating Electron Transfer Flavoprotein (BfETF), we are assessing the applicability of several possible mechanisms that could bar the high-energy electron from exploiting the exergonic path and dissipating energy. Our work demonstrates that thermodynamic considerations may prevent a second electron from using to the endergonic path, but this presentation will focus on possible significance of an enormous domain-scale conformational change that is documented in structural studies and proposed to gate electron transfer.
Accelerated molecular dynamics (MD) trajectories and quantum mechanical (QM) calculations are underway to understand what motions may be coupled to catalytic events (as opposed to reflecting fluctuations inherent in the protein structure). These approaches enable us to assess the significance of redox-coupled movements of individual protons and reorganization of crucial hydrogen bond networks in the interfaces between domains proposed to move relative to one-another. Meanwhile, we are using Small Angle Neutron Scattering (SANS) and Nuclear Magnetic Resonance (NMR) to learn what conformations may be present under conditions of catalytic activity. SANS and NMR are ideally suited to the task because they do not perturb the oxidation states of flavins, which constitute key steps in BfETF turnover. Unfortunately, the common X-ray and FRET-related methods can cause photoreduction of flavins and therefore could trigger the conformational change we seek to control and characterize. Indeed, our new data reveal subtle but consistent changes in the SANS profiles in response to reduction of the BfETF. Thus we have evidence that a conformational event is indeed coupled to step(s) in bifurcation, in BfETF.
A.-F. M. is pleased to acknowledge partial support from D.O.E. DESC0021283, KY-EPSCoR PON2 635 2000003148 for investigations of mechanisms for coupling conformational change to flavin-redox events, and from N.S.F. CHE 2108134 for investigation of H-bonding as a mechanism tuning flavin reduction midpoint potentials. S.A.K. acknowledges partial support from the Oak Ridge National Laboratory (ORNL) neutron scattering GRO program. We thank the Einstein Foundation of Berlin for a visiting fellowship to AFM, and ongoing support to MGV via the Einstein Center for Catalysis. MAM's thanks the DFG for funding under SFB1078, project C2. SANS studies were performed using Bio-SANS instrument of the Center for Structural Molecular Biology (FWP ERKP291) and a DOE Office of Biological and Environmental Research Structural Biology Resource. This research used resources at the High Flux Isotope Reactor and Spallation Neutron Source, a DOE Office of Science User Facility operated by the ORNL. ORNL is operated by UT-Battelle, LLC under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy.
| Topic | Biological Energy Transfer |
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