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TitleCryo-EM structures define the electron bifurcating flavobicluster and ferredoxin binding site in an archaeal Nfn-Bfu transhydrogenase.
Journal, issue, pagesJ Biol Chem, Vol. 301, Issue 4, Page 108410, Year 2025
Publish dateMar 18, 2025
AuthorsXiansha Xiao / Gerrit J Schut / Xiang Feng / Diep M N Nguyen / Haiyan Huang / Shuning Wang / Huilin Li / Michael W W Adams /
PubMed AbstractFlavin-based electron bifurcation couples exergonic and endergonic redox reactions in one enzyme complex to circumvent thermodynamic barriers and minimize free energy loss. Two unrelated enzymes ...Flavin-based electron bifurcation couples exergonic and endergonic redox reactions in one enzyme complex to circumvent thermodynamic barriers and minimize free energy loss. Two unrelated enzymes designated NfnSL and NfnABC catalyze the NADPH-dependent reduction of ferredoxin and NAD. Bifurcation by NfnSL resides with a single FAD but the bifurcation mechanism of NfnABC, which represents the diverse and ubiquitous Bfu enzyme family, is completely different and largely unknown. Using cryo-EM structures of an archaeal NfnABC, we show that its bifurcation site is a flavobicluster consisting of FMN, one [4Fe-4S] and one [2Fe-2S] cluster where zinc atoms replace two additional clusters previously identified in other Bfu enzymes. NADH binds to the flavobicluster site of NfnABC and induces conformational changes that allow ferredoxin to bind between the C-terminal domains of NfnC and NfnB. Site-directed mutational analyses support the proposed mechanism that is likely conserved in all members of the Bfu enzyme family.
External linksJ Biol Chem / PubMed:40107619 / PubMed Central
MethodsEM (single particle)
Resolution2.76 - 2.85 Å
Structure data

49039

9n5u

EMDB-49039: Cryo-EM map of the Thermococcus sibiricus NfnABC complex
PDB-9n5u: Structure of the Thermococcus sibiricus NfnABC complex
Method: EM (single particle) / Resolution: 2.85 Å

49040

9n5v

EMDB-49040: Cryo-EM map of the NAD(H)-bound Thermococcus sibiricus NfnABC complex
PDB-9n5v: Structure of the NAD(H)-bound Thermococcus sibiricus NfnABC complex
Method: EM (single particle) / Resolution: 2.76 Å

Chemicals

ChemComp-SF4:
IRON/SULFUR CLUSTER

ChemComp-FAD:
FLAVIN-ADENINE DINUCLEOTIDE / FAD*YM

ChemComp-FES:
FE2/S2 (INORGANIC) CLUSTER

ChemComp-FMN:
FLAVIN MONONUCLEOTIDE

ChemComp-ZN:
Unknown entry

ChemComp-NAD:
NICOTINAMIDE-ADENINE-DINUCLEOTIDE / NAD*YM

Source
  • thermococcus sibiricus (archaea)
KeywordsELECTRON TRANSPORT / Flavin-based electron bifurcation / Bfu family / Nfn-ABC / cryo-EM structures / flavo-FeS cluster / NfnABC

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