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- PDB-9erl: Cryo-EM structure of sodium pumping Rnf complex from Acetobacteri... -

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Basic information

Entry
Database: PDB / ID: 9erl
TitleCryo-EM structure of sodium pumping Rnf complex from Acetobacterium woodii in apo state
Components(Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit ...) x 6
KeywordsMEMBRANE PROTEIN / Bioenergetics / anaerobic cryoEM / electron transport / redox-driven sodium pumping
Function / homology
Function and homology information


ferredoxin-NAD+ oxidoreductase (Na+-transporting) / electron transport chain / transmembrane transport / FMN binding / 4 iron, 4 sulfur cluster binding / electron transfer activity / metal ion binding / plasma membrane
Similarity search - Function
Ion-translocating oxidoreductase complex, RnfB/RsxB / Ion-translocating oxidoreductase complex, subunit RnfC/RsxC / Ion-translocating oxidoreductase complex subunit E / Ion-translocating oxidoreductase complex, subunit RnfA/RsxA / Ion-translocating oxidoreductase complex Rnf, subunit D / RnfC Barrel sandwich hybrid domain / : / RnfC Barrel sandwich hybrid domain / Ion-translocating oxidoreductase complex, subunit RnfG/RsxG / 4Fe-4S domain ...Ion-translocating oxidoreductase complex, RnfB/RsxB / Ion-translocating oxidoreductase complex, subunit RnfC/RsxC / Ion-translocating oxidoreductase complex subunit E / Ion-translocating oxidoreductase complex, subunit RnfA/RsxA / Ion-translocating oxidoreductase complex Rnf, subunit D / RnfC Barrel sandwich hybrid domain / : / RnfC Barrel sandwich hybrid domain / Ion-translocating oxidoreductase complex, subunit RnfG/RsxG / 4Fe-4S domain / Putative Fe-S cluster / 4Fe-4S domain profile. / NqrDE/RnfAE / Ion-translocating oxidoreductase NqrB/RnfD / : / Rnf-Nqr subunit, membrane protein / NQR2, RnfD, RnfE family / FMN-binding / FMN-binding domain / FMN_bind / 4Fe-4S dicluster domain / Soluble ligand binding domain / SLBB domain / 4Fe-4S binding domain / NADH-ubiquinone oxidoreductase 51kDa subunit, FMN-binding domain / NADH-ubiquinone oxidoreductase 51kDa subunit, FMN-binding domain superfamily / Nuo51 FMN-binding domain / 4Fe-4S dicluster domain / 4Fe-4S ferredoxin, iron-sulphur binding, conserved site / 4Fe-4S ferredoxin-type iron-sulfur binding region signature. / 4Fe-4S ferredoxin-type iron-sulfur binding domain profile. / 4Fe-4S ferredoxin-type, iron-sulphur binding domain
Similarity search - Domain/homology
FE2/S2 (INORGANIC) CLUSTER / FLAVIN MONONUCLEOTIDE / RIBOFLAVIN / IRON/SULFUR CLUSTER / Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit B / Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit A / Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit E / Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit G / Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit D / Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit C
Similarity search - Component
Biological speciesAcetobacterium woodii DSM 1030 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å
AuthorsKumar, A. / Schuller, J.M.
Funding support Germany, European Union, 2items
OrganizationGrant numberCountry
German Research Foundation (DFG)SCHU 3364/1-1 Germany
European Research Council (ERC)101075992European Union
CitationJournal: Nat Commun / Year: 2025
Title: Molecular principles of redox-coupled sodium pumping of the ancient Rnf machinery.
Authors: Anuj Kumar / Jennifer Roth / Hyunho Kim / Patricia Saura / Stefan Bohn / Tristan Reif-Trauttmansdorff / Anja Schubert / Ville R I Kaila / Jan M Schuller / Volker Müller /
Abstract: The Rnf complex is the primary respiratory enzyme of several anaerobic prokaryotes that transfers electrons from ferredoxin to NAD and pumps ions (Na or H) across a membrane, powering ATP synthesis. ...The Rnf complex is the primary respiratory enzyme of several anaerobic prokaryotes that transfers electrons from ferredoxin to NAD and pumps ions (Na or H) across a membrane, powering ATP synthesis. Rnf is widespread in primordial organisms and the evolutionary predecessor of the Na-pumping NADH-quinone oxidoreductase (Nqr). By running in reverse, Rnf uses the electrochemical ion gradient to drive ferredoxin reduction with NADH, providing low potential electrons for nitrogenases and CO reductases. Yet, the molecular principles that couple the long-range electron transfer to Na translocation remain elusive. Here, we resolve key functional states along the electron transfer pathway in the Na-pumping Rnf complex from Acetobacterium woodii using redox-controlled cryo-electron microscopy that, in combination with biochemical functional assays and atomistic molecular simulations, provide key insight into the redox-driven Na pumping mechanism. We show that the reduction of the unique membrane-embedded [2Fe2S] cluster electrostatically attracts Na, and in turn, triggers an inward/outward transition with alternating membrane access driving the Na pump and the reduction of NAD. Our study unveils an ancient mechanism for redox-driven ion pumping, and provides key understanding of the fundamental principles governing energy conversion in biological systems.
History
DepositionMar 23, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 5, 2025Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: Additional map / Part number: 1 / Data content type: Additional map / Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.1Mar 19, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit A
B: Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit B
C: Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit C
D: Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit D
E: Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit E
G: Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit G
hetero molecules


Theoretical massNumber of molelcules
Total (without water)183,98924
Polymers178,4836
Non-polymers5,50718
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area26060 Å2
ΔGint-467 kcal/mol
Surface area75230 Å2
MethodPISA

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Components

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Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit ... , 6 types, 6 molecules ABCDEG

#1: Protein Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit A / Rnf electron transport complex subunit A


Mass: 20384.445 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Acetobacterium woodii DSM 1030 (bacteria)
References: UniProt: H6LC28, ferredoxin-NAD+ oxidoreductase (Na+-transporting)
#2: Protein Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit B / Rnf electron transport complex subunit B


Mass: 34762.801 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Acetobacterium woodii DSM 1030 (bacteria)
References: UniProt: H6LC27, ferredoxin-NAD+ oxidoreductase (Na+-transporting)
#3: Protein Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit C / Rnf electron transport complex subunit C


Mass: 47177.758 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Acetobacterium woodii DSM 1030 (bacteria)
References: UniProt: H6LC32, ferredoxin-NAD+ oxidoreductase (Na+-transporting)
#4: Protein Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit D / Rnf electron transport complex subunit D


Mass: 33762.004 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Acetobacterium woodii DSM 1030 (bacteria)
References: UniProt: H6LC31, ferredoxin-NAD+ oxidoreductase (Na+-transporting)
#5: Protein Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit E / Rnf electron transport complex subunit E


Mass: 20544.832 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Acetobacterium woodii DSM 1030 (bacteria)
References: UniProt: H6LC29, ferredoxin-NAD+ oxidoreductase (Na+-transporting)
#6: Protein Na(+)-translocating ferredoxin:NAD(+) oxidoreductase complex subunit G / Rnf electron transport complex subunit G


Mass: 21850.881 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Acetobacterium woodii DSM 1030 (bacteria)
References: UniProt: H6LC30, ferredoxin-NAD+ oxidoreductase (Na+-transporting)

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Non-polymers , 5 types, 18 molecules

#7: Chemical ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Na / Feature type: SUBJECT OF INVESTIGATION
#8: Chemical ChemComp-FES / FE2/S2 (INORGANIC) CLUSTER


Mass: 175.820 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Fe2S2 / Feature type: SUBJECT OF INVESTIGATION
#9: Chemical
ChemComp-SF4 / IRON/SULFUR CLUSTER


Mass: 351.640 Da / Num. of mol.: 10 / Source method: obtained synthetically / Formula: Fe4S4 / Feature type: SUBJECT OF INVESTIGATION
#10: Chemical ChemComp-FMN / FLAVIN MONONUCLEOTIDE / RIBOFLAVIN MONOPHOSPHATE


Mass: 456.344 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C17H21N4O9P / Feature type: SUBJECT OF INVESTIGATION
#11: Chemical ChemComp-RBF / RIBOFLAVIN / RIBOFLAVINE / VITAMIN B2


Mass: 376.364 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C17H20N4O6 / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: rhodobacter nitrogen fixation complex from Acetobacterium woodii
Type: COMPLEX / Entity ID: #1-#6 / Source: NATURAL
Molecular weightValue: 0.16 MDa / Experimental value: YES
Source (natural)Organism: Acetobacterium woodii DSM 1030 (bacteria)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: The Rnf complex was reduced with low potential ferredoxin.
VitrificationCryogen name: ETHANE-PROPANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2200 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 251476 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00312922
ELECTRON MICROSCOPYf_angle_d0.65217628
ELECTRON MICROSCOPYf_dihedral_angle_d5.2811861
ELECTRON MICROSCOPYf_chiral_restr0.0452140
ELECTRON MICROSCOPYf_plane_restr0.0072169

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