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- PDB-8s7x: Methyl-coenzyme M reductase activation complex without the A2 com... -

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Basic information

Entry
Database: PDB / ID: 8s7x
TitleMethyl-coenzyme M reductase activation complex without the A2 component
Components
  • (Methanogenesis marker protein ...) x 2
  • (Methyl-coenzyme M reductase subunit ...) x 3
  • DUF2098 domain-containing protein
  • Methyl-coenzyme M reductase operon protein C
  • UPF0288 protein MmarC6_0796
KeywordsOXIDOREDUCTASE / Methyl-coenzyme M reductase / activation complex / Iron-sulfur clusters
Function / homology
Function and homology information


coenzyme-B sulfoethylthiotransferase / coenzyme-B sulfoethylthiotransferase activity / methanogenesis / metal ion binding / cytoplasm
Similarity search - Function
Methyl-coenzyme M reductase, protein C / Uncharacterised conserved protein UCP019164, methanogenesis / Uncharacterised protein family UPF0288, methanogenesis / Uncharacterised conserved protein UCP019464, methanogenesis / Uncharacterised conserved protein UCP037053 / Protein of unknown function DUF2098 / Methyl-coenzyme M reductase, protein C-like / Methyl-coenzyme M reductase operon protein C / Uncharacterized protein conserved in archaea (DUF2098) / Uncharacterized protein conserved in archaea (DUF2113) ...Methyl-coenzyme M reductase, protein C / Uncharacterised conserved protein UCP019164, methanogenesis / Uncharacterised protein family UPF0288, methanogenesis / Uncharacterised conserved protein UCP019464, methanogenesis / Uncharacterised conserved protein UCP037053 / Protein of unknown function DUF2098 / Methyl-coenzyme M reductase, protein C-like / Methyl-coenzyme M reductase operon protein C / Uncharacterized protein conserved in archaea (DUF2098) / Uncharacterized protein conserved in archaea (DUF2113) / Methyl-coenzyme M reductase, alpha subunit, N-terminal subdomain 1 / Methyl-coenzyme M reductase, gamma subunit / Methyl-coenzyme M reductase, beta subunit / Methyl coenzyme M reductase, alpha subunit / Methyl-coenzyme M reductase, beta subunit, C-terminal / Methyl-coenzyme M reductase, beta subunit, N-terminal / Methyl-coenzyme M reductase, gamma subunit superfamily / Methyl-coenzyme M reductase gamma subunit / Methyl-coenzyme M reductase beta subunit, C-terminal domain / Methyl-coenzyme M reductase beta subunit, N-terminal domain / Methyl-coenzyme M reductase, alpha subunit, N-terminal / Methyl-coenzyme M reductase, alpha/beta subunit, C-terminal / Methyl-coenzyme M reductase, ferredoxin-like fold / Methyl-coenzyme M reductase, alpha subunit, C-terminal / Methyl-coenzyme M reductase, alpha subunit, N-terminal subdomain 2 / Methyl-coenzyme M reductase alpha subunit, C-terminal domain / Methyl-coenzyme M reductase alpha subunit, N-terminal domain
Similarity search - Domain/homology
1-THIOETHANESULFONIC ACID / FACTOR 430 / FeFe cofactor / Chem-SHT / Coenzyme B / DUF2098 domain-containing protein / Methyl-coenzyme M reductase subunit alpha / Methyl-coenzyme M reductase subunit beta / Methyl-coenzyme M reductase subunit gamma / UPF0288 protein MmarC6_0796 ...1-THIOETHANESULFONIC ACID / FACTOR 430 / FeFe cofactor / Chem-SHT / Coenzyme B / DUF2098 domain-containing protein / Methyl-coenzyme M reductase subunit alpha / Methyl-coenzyme M reductase subunit beta / Methyl-coenzyme M reductase subunit gamma / UPF0288 protein MmarC6_0796 / Methyl-coenzyme M reductase operon protein C / Methanogenesis marker protein 17 / Methanogenesis marker protein 7
Similarity search - Component
Biological speciesMethanococcus maripaludis (archaea)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.78 Å
AuthorsRamirez-Amador, F. / Paul, S. / Kumar, A. / Schuller, J.M.
Funding supportEuropean Union, 1items
OrganizationGrant numberCountry
European Research Council (ERC)101075992European Union
CitationJournal: Nature / Year: 2025
Title: Structure of the ATP-driven methyl-coenzyme M reductase activation complex.
Authors: Fidel Ramírez-Amador / Sophia Paul / Anuj Kumar / Christian Lorent / Sebastian Keller / Stefan Bohn / Thinh Nguyen / Stefano Lometto / Dennis Vlegels / Jörg Kahnt / Darja Deobald / Frank ...Authors: Fidel Ramírez-Amador / Sophia Paul / Anuj Kumar / Christian Lorent / Sebastian Keller / Stefan Bohn / Thinh Nguyen / Stefano Lometto / Dennis Vlegels / Jörg Kahnt / Darja Deobald / Frank Abendroth / Olalla Vázquez / Georg Hochberg / Silvan Scheller / Sven T Stripp / Jan Michael Schuller /
Abstract: Methyl-coenzyme M reductase (MCR) is the enzyme responsible for nearly all biologically generated methane. Its active site comprises coenzyme F, a porphyrin-based cofactor with a central nickel ion ...Methyl-coenzyme M reductase (MCR) is the enzyme responsible for nearly all biologically generated methane. Its active site comprises coenzyme F, a porphyrin-based cofactor with a central nickel ion that is active exclusively in the Ni(I) state. How methanogenic archaea perform the reductive activation of F represents a major gap in our understanding of one of the most ancient bioenergetic systems in nature. Here we purified and characterized the MCR activation complex from Methanococcus maripaludis. McrC, a small subunit encoded in the mcr operon, co-purifies with the methanogenic marker proteins Mmp7, Mmp17, Mmp3 and the A2 component. We demonstrated that this complex can activate MCR in vitro in a strictly ATP-dependent manner, enabling the formation of methane. In addition, we determined the cryo-electron microscopy structure of the MCR activation complex exhibiting different functional states with local resolutions reaching 1.8-2.1 Å. Our data revealed three complex iron-sulfur clusters that formed an electron transfer pathway towards F. Topology and electron paramagnetic resonance spectroscopy analyses indicate that these clusters are similar to the [8Fe-9S-C] cluster, a maturation intermediate of the catalytic cofactor in nitrogenase. Altogether, our findings offer insights into the activation mechanism of MCR and prospects on the early evolution of nitrogenase.
History
DepositionMar 4, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 26, 2025Provider: repository / Type: Initial release
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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Methyl-coenzyme M reductase subunit gamma
B: Methyl-coenzyme M reductase subunit gamma
E: Methyl-coenzyme M reductase subunit beta
D: Methyl-coenzyme M reductase subunit beta
C: Methyl-coenzyme M reductase subunit alpha
G: Methanogenesis marker protein 17
F: Methyl-coenzyme M reductase subunit alpha
H: Methanogenesis marker protein 7
I: Methyl-coenzyme M reductase operon protein C
J: UPF0288 protein MmarC6_0796
L: DUF2098 domain-containing protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)428,46219
Polymers423,43911
Non-polymers5,0228
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Methyl-coenzyme M reductase subunit ... , 3 types, 6 molecules ABEDCF

#1: Protein Methyl-coenzyme M reductase subunit gamma


Mass: 29665.488 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Details: Methyl-coenzyme M reductase subunit gamma / Source: (natural) Methanococcus maripaludis (archaea) / Strain: JJ -upt
References: UniProt: A0A2L1CBG2, coenzyme-B sulfoethylthiotransferase
#2: Protein Methyl-coenzyme M reductase subunit beta / Coenzyme-B sulfoethylthiotransferase beta


Mass: 46731.230 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Details: Methyl-coenzyme M reductase subunit beta / Source: (natural) Methanococcus maripaludis (archaea) / Strain: JJ -upt
References: UniProt: A0A2L1CBB3, coenzyme-B sulfoethylthiotransferase
#3: Protein Methyl-coenzyme M reductase subunit alpha


Mass: 61230.703 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Details: Methyl-coenzyme M reductase subunit alpha / Source: (natural) Methanococcus maripaludis (archaea) / Strain: JJ -upt
References: UniProt: A0A2L1CBB0, coenzyme-B sulfoethylthiotransferase

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Methanogenesis marker protein ... , 2 types, 2 molecules GH

#4: Protein Methanogenesis marker protein 17 / Mmp17


Mass: 21119.539 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: Methanogenesis marker protein 17 / Source: (natural) Methanococcus maripaludis (archaea) / Strain: JJ -upt / References: UniProt: G0H411
#5: Protein Methanogenesis marker protein 7


Mass: 35024.746 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: Methanogenesis marker protein 7 / Source: (natural) Methanococcus maripaludis (archaea) / Strain: JJ -upt / References: UniProt: Q6M050

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Protein , 3 types, 3 molecules IJL

#6: Protein Methyl-coenzyme M reductase operon protein C


Mass: 24906.715 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: N-terminally Twin-Strep-tagged Methyl-coenzyme M reductase operon protein C
Source: (gene. exp.) Methanococcus maripaludis (archaea) / Strain: JJ -upt / Gene: GYY_08635 / Production host: Methanococcus maripaludis (archaea) / Strain (production host): JJ -upt / References: UniProt: G0H3B1
#7: Protein UPF0288 protein MmarC6_0796 / Methanogenesis marker protein 3 / Mmp3


Mass: 56520.801 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: Methanogenesis marker protein 3 / Source: (natural) Methanococcus maripaludis (archaea) / Strain: JJ -upt / References: UniProt: A9A8E0
#8: Protein DUF2098 domain-containing protein


Mass: 10612.835 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: Domain of unknown function protein 2098 / Source: (natural) Methanococcus maripaludis (archaea) / Strain: JJ -upt / References: UniProt: A0A2L1CAX0

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Non-polymers , 5 types, 8 molecules

#9: Chemical ChemComp-F43 / FACTOR 430


Mass: 906.580 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C42H51N6NiO13
#10: Chemical ChemComp-SHT / O-PHOSPHONO-N-{(2E)-7-[(2-SULFOETHYL)DITHIO]HEPT-2-ENOYL}-L-THREONINE


Mass: 481.499 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C13H24NO10PS3 / Feature type: SUBJECT OF INVESTIGATION
#11: Chemical ChemComp-COM / 1-THIOETHANESULFONIC ACID


Mass: 142.197 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C2H6O3S2
#12: Chemical ChemComp-TP7 / Coenzyme B / 7-MERCAPTOHEPTANOYLTHREONINEPHOSPHATE


Mass: 343.334 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C11H22NO7PS
#13: Chemical ChemComp-S5Q / FeFe cofactor


Mass: 747.356 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: CFe8S9 / 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: 3D ARRAY / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Methyl-coenzyme M reductase activation complex without A2 component
Type: COMPLEX / Entity ID: #1, #3, #2, #4, #7-#8 / Source: NATURAL
Molecular weightValue: 0.42 MDa / Experimental value: NO
Source (natural)Organism: Methanococcus maripaludis (archaea)
Buffer solutionpH: 7.6
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
2150 mMsodium chlorideNaCl1
SpecimenConc.: 1.25 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 15 mA / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277 K
Details: Vitrification setup inside a Coy Lab's Vinyl Anaerobic Chamber to preserve the protein sample under strict anaerobic conditions (95% O2 / 5% H2).

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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 magnification: 165000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k) / Num. of real images: 17548
Details: 7781 micrographs with a 20 deg pretilt to overcome preferred orientation problems

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Processing

EM software
IDNameVersionCategory
1cryoSPARCv4particle selection
2cryoSPARCv4image acquisition
4cryoSPARCCTF correction
7UCSF Chimera1.17.3model fitting
8Cootmodel fitting
10PHENIXmodel refinement
13cryoSPARCclassification
14cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1557902
Details: 1023667 from non tilted and 534235 from pretilted datasets
3D reconstructionResolution: 2.78 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 349437 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
Details: Rigid body fitting in Chimera and Coot. Further real-space refinement in Phenix.
Atomic model buildingSource name: AlphaFold / Type: in silico model

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