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- EMDB-2096: Cryo-EM structure of the F420-reducing [NiFe]-hydrogenase from a ... -

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Basic information

Entry
Database: EMDB / ID: EMD-2096
TitleCryo-EM structure of the F420-reducing [NiFe]-hydrogenase from a methanogenic archaeon at near-atomic resolution
Map datareconstruction of Frh complex
Sample
  • Sample: F420-reducing [NiFe] hydrogenase Frh
  • Protein or peptide: F420-reducing [NiFe] hydrogenase
Keywordshydrogenase / [NiFe] hydrogenase / methanogenesis / F420-reducing hydrogenase
Function / homology
Function and homology information


coenzyme F420 hydrogenase / coenzyme F420 hydrogenase activity / oxidoreductase activity, acting on CH or CH2 groups, with an iron-sulfur protein as acceptor / ferredoxin hydrogenase activity / iron-sulfur cluster binding / nickel cation binding / flavin adenine dinucleotide binding / 4 iron, 4 sulfur cluster binding
Similarity search - Function
Coenzyme F420 hydrogenase, subunit alpha / Coenzyme F420 hydrogenase, subunit alpha / Coenzyme F420 hydrogenase subunit beta, archaea / Coenzyme F420 hydrogenase, subunit gamma / Coenzyme F420 hydrogenase subunit beta, archaea / Coenzyme F420 hydrogenase, subunit gamma / : / Coenzyme F420 hydrogenase/dehydrogenase beta subunit, N-terminal / Coenzyme F420 hydrogenase/dehydrogenase beta subunit, C-terminal / Oxidoreductase FRHB/FDHB/HCAR-like ...Coenzyme F420 hydrogenase, subunit alpha / Coenzyme F420 hydrogenase, subunit alpha / Coenzyme F420 hydrogenase subunit beta, archaea / Coenzyme F420 hydrogenase, subunit gamma / Coenzyme F420 hydrogenase subunit beta, archaea / Coenzyme F420 hydrogenase, subunit gamma / : / Coenzyme F420 hydrogenase/dehydrogenase beta subunit, N-terminal / Coenzyme F420 hydrogenase/dehydrogenase beta subunit, C-terminal / Oxidoreductase FRHB/FDHB/HCAR-like / Coenzyme F420 hydrogenase/dehydrogenase, beta subunit N-term / Coenzyme F420 hydrogenase/dehydrogenase, beta subunit C terminus / Nickel-dependent hydrogenases large subunit signature 2. / Nickel-dependent hydrogenases large subunit signature 1. / [NiFe]-hydrogenase, small subunit, N-terminal domain superfamily / Nickel-dependent hydrogenase, large subunit, nickel binding site / Nickel-dependent hydrogenase, large subunit / Nickel-dependent hydrogenase / 4Fe-4S binding domain / NADH:ubiquinone oxidoreductase-like, 20kDa subunit / NADH ubiquinone oxidoreductase, 20 Kd subunit / [NiFe]-hydrogenase, large subunit / 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
Coenzyme F420 hydrogenase subunit beta / Coenzyme F420 hydrogenase subunit gamma / Coenzyme F420 hydrogenase subunit alpha
Similarity search - Component
Biological speciesMethanothermobacter marburgensis (archaea)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.5 Å
AuthorsMills DJ / Vitt S / Strauss M / Shima S / Vonck J
CitationJournal: Elife / Year: 2013
Title: De novo modeling of the F(420)-reducing [NiFe]-hydrogenase from a methanogenic archaeon by cryo-electron microscopy.
Authors: Deryck J Mills / Stella Vitt / Mike Strauss / Seigo Shima / Janet Vonck /
Abstract: Methanogenic archaea use a [NiFe]-hydrogenase, Frh, for oxidation/reduction of F420, an important hydride carrier in the methanogenesis pathway from H2 and CO2. Frh accounts for about 1% of the ...Methanogenic archaea use a [NiFe]-hydrogenase, Frh, for oxidation/reduction of F420, an important hydride carrier in the methanogenesis pathway from H2 and CO2. Frh accounts for about 1% of the cytoplasmic protein and forms a huge complex consisting of FrhABG heterotrimers with each a [NiFe] center, four Fe-S clusters and an FAD. Here, we report the structure determined by near-atomic resolution cryo-EM of Frh with and without bound substrate F420. The polypeptide chains of FrhB, for which there was no homolog, was traced de novo from the EM map. The 1.2-MDa complex contains 12 copies of the heterotrimer, which unexpectedly form a spherical protein shell with a hollow core. The cryo-EM map reveals strong electron density of the chains of metal clusters running parallel to the protein shell, and the F420-binding site is located at the end of the chain near the outside of the spherical structure. DOI:http://dx.doi.org/10.7554/eLife.00218.001.
History
DepositionMay 10, 2012-
Header (metadata) releaseMay 25, 2012-
Map releaseMar 13, 2013-
UpdateJul 3, 2013-
Current statusJul 3, 2013Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 212
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 212
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-3zfs
  • Surface level: 212
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-3zfs
  • Surface level: 212
  • Imaged by UCSF Chimera
  • Download
  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-3zfs
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

EMD-2096.jpg

Downloads & links

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Map

FileDownload / File: emd_2096.map.gz / Format: CCP4 / Size: 89 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationreconstruction of Frh complex
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.14 Å/pix.
x 288 pix.
= 328.32 Å		1.14 Å/pix.
x 288 pix.
= 328.32 Å		1.14 Å/pix.
x 288 pix.
= 328.32 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.14 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 212.0 / Movie #1: 212
Minimum - Maximum-506.050567629999989 - 1455.878051760000062
Average (Standard dev.)3.10870266 (±46.902236940000002)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 328.32 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.141.141.14
M x/y/z288288288
origin x/y/z0.0000.0000.000
length x/y/z328.320328.320328.320
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ128128168
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS288288288
D min/max/mean-506.0511455.8783.109

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Supplemental data

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Sample components

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Entire : F420-reducing [NiFe] hydrogenase Frh

EntireName: F420-reducing [NiFe] hydrogenase Frh
Components
  • Sample: F420-reducing [NiFe] hydrogenase Frh
  • Protein or peptide: F420-reducing [NiFe] hydrogenase

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Supramolecule #1000: F420-reducing [NiFe] hydrogenase Frh

SupramoleculeName: F420-reducing [NiFe] hydrogenase Frh / type: sample / ID: 1000 / Oligomeric state: dodecamer / Number unique components: 1
Molecular weightExperimental: 1.2 MDa / Theoretical: 1.2 MDa

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Macromolecule #1: F420-reducing [NiFe] hydrogenase

MacromoleculeName: F420-reducing [NiFe] hydrogenase / type: protein_or_peptide / ID: 1 / Name.synonym: Frh, FrhABG
Details: The Frh complex is a tetrahedral dodecamer of an FrhA, FrhB, FrhG heterotrimer
Number of copies: 12 / Oligomeric state: dodecamer / Recombinant expression: No
Source (natural)Organism: Methanothermobacter marburgensis (archaea) / Strain: DSM 2133 / Location in cell: cytoplasm
Molecular weightExperimental: 1.2 MDa / Theoretical: 1.2 MDa
SequenceInterPro: Coenzyme F420 hydrogenase, subunit alpha, Coenzyme F420 hydrogenase subunit beta, archaea, Coenzyme F420 hydrogenase, subunit gamma

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.7 mg/mL
BufferpH: 7.6 / Details: 50mM Tris-HCl, 0.025mM FAD
GridDetails: freshly glow discharged Quantifoil holey grid (1 micrometer holes)
VitrificationCryogen name: ETHANE / Chamber humidity: 70 % / Chamber temperature: 103 K / Instrument: FEI VITROBOT MARK I / Method: blotting 2.5 seconds before plunging

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Electron microscopy

MicroscopeFEI POLARA 300
TemperatureMin: 77 K / Max: 80 K / Average: 79 K
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 120,000 times magnification
DateAug 26, 2010
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Number real images: 101 / Average electron dose: 15 e/Å2 / Bits/pixel: 8
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 61400 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 2.8 µm / Nominal defocus min: 1.69 µm / Nominal magnification: 59000
Sample stageSpecimen holder model: OTHER
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

DetailsEMAN2
CTF correctionDetails: each micrograph
Final reconstructionApplied symmetry - Point group: T (tetrahedral) / Resolution.type: BY AUTHOR / Resolution: 4.5 Å / Resolution method: OTHER / Software - Name: EMAN2 / Number images used: 84000
Final two d classificationNumber classes: 2134

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Atomic model buiding 1

Initial modelPDB ID:

2wpn


Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B
SoftwareName: chimera, coot
DetailsProtocol: A rigid body fit was done in Chimera and the model manually rebuilt in Coot.
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-3zfs:
Cryo-EM structure of the F420-reducing NiFe-hydrogenase from a methanogenic archaeon with bound substrate

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