Abstract

Bifurcating electron transferring flavoproteins (Bf-ETFs) tune chemically identical flavins to two contrasting roles. To understand how, we used hybrid quantum mechanical molecular mechanical calculations to characterize noncovalent interactions applied to each flavin by the protein. Our computations replicated the differences between the reactivities of the flavins: the electron transferring flavin ( ETflavin) was calculated to stabilize anionic semiquinone (ASQ) as needed to execute its single-electron transfers, whereas the Bf flavin (Bfflavin) was found to disfavor the ASQ state more than does free flavin and to be less susceptible to reduction. The stability of ETflavin ASQ was attributed in part to H-bond donation to the flavin O2 from a nearby His side chain, via comparison of models employing different tautomers of His. This H-bond between O2 and the ET site was uniquely strong in the ASQ state, whereas reduction of ETflavin to the anionic hydroquinone (AHQ) was associated with side chain reorientation, backbone displacement, and reorganization of its H-bond network including a Tyr from the other domain and subunit of the ETF. The Bf site was less responsive overall, but formation of the Bfflavin AHQ allowed a nearby Arg side chain to adopt an alternative rotamer that can H-bond to the Bfflavin O4. This would stabilize the anionic Bfflavin and rationalize effects of mutation at this position. Thus, our computations provide insights on states and conformations that have not been possible to characterize experimentally, offering explanations for observed residue conservation and raising possibilities that can now be tested.

Document Type

Article

Publication Date

6-2023

Notes/Citation Information

© 2023 THE AUTHORS. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Digital Object Identifier (DOI)

https://doi.org/10.1016/j.jbc.2023.104762

Funding Information

A.-F. M. is pleased to acknowledge partial support from D.O.E. DESC0021283 and 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 investigations of H-bonding as a mechanism tuning flavin reduction midpoint potentials. A.-F. M. acknowledges WDW for vital assistance, and RNM for perseverance. We thank the Einstein Foundation of Berlin for funding to R. K. K., a visiting fellowship to A.-F. M., and ongoing support to M. G.-V. via the Einstein Center for Catalysis. M.-A. M.’s research was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under SFB1078, project C2. R. K. K. thanks DST-SERB for a start-up grant (SRG/2022/000858).

Download

Share

COinS