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- PDB-8dqv: The 1.52 angstrom CryoEM structure of the [NiFe]-hydrogenase Huc ... -

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Basic information

Entry
Database: PDB / ID: 8dqv
TitleThe 1.52 angstrom CryoEM structure of the [NiFe]-hydrogenase Huc from Mycobacterium smegmatis - catalytic dimer (Huc2S2L)
Components(Hydrogenase-2, ...) x 2
KeywordsELECTRON TRANSPORT / [NiFe] Hydrogenase / Membrane-associated / Complex / Quinone Transport
Function / homology
Function and homology information


hydrogenase (acceptor) / ferredoxin hydrogenase complex / hydrogenase (acceptor) activity / ferredoxin hydrogenase activity / 3 iron, 4 sulfur cluster binding / nickel cation binding / 4 iron, 4 sulfur cluster binding / metal ion binding
Similarity search - Function
[NiFe]-hydrogenase, small subunit / Cytochrome-c3 hydrogenase, C-terminal / [NiFe]-hydrogenase, small subunit, C-terminal domain superfamily / NiFe/NiFeSe hydrogenase small subunit C-terminal / 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 / NADH:ubiquinone oxidoreductase-like, 20kDa subunit ...[NiFe]-hydrogenase, small subunit / Cytochrome-c3 hydrogenase, C-terminal / [NiFe]-hydrogenase, small subunit, C-terminal domain superfamily / NiFe/NiFeSe hydrogenase small subunit C-terminal / 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 / NADH:ubiquinone oxidoreductase-like, 20kDa subunit / NADH ubiquinone oxidoreductase, 20 Kd subunit / [NiFe]-hydrogenase, large subunit
Similarity search - Domain/homology
NICKEL (III) ION / FE3-S4 CLUSTER / CARBONMONOXIDE-(DICYANO) IRON / OXYGEN ATOM / MENADIONE / Hydrogenase-2, large subunit / NADH ubiquinone oxidoreductase 20 kDa subunit
Similarity search - Component
Biological speciesMycolicibacterium smegmatis (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 1.52 Å
AuthorsGrinter, R. / Venugopal, H. / Kropp, A. / Greening, C.
Funding support Australia, 2items
OrganizationGrant numberCountry
Australian Research Council (ARC)DP200103074 Australia
National Health and Medical Research Council (NHMRC, Australia)APP1197376 Australia
CitationJournal: Nature / Year: 2023
Title: Structural basis for bacterial energy extraction from atmospheric hydrogen.
Authors: Rhys Grinter / Ashleigh Kropp / Hari Venugopal / Moritz Senger / Jack Badley / Princess R Cabotaje / Ruyu Jia / Zehui Duan / Ping Huang / Sven T Stripp / Christopher K Barlow / Matthew ...Authors: Rhys Grinter / Ashleigh Kropp / Hari Venugopal / Moritz Senger / Jack Badley / Princess R Cabotaje / Ruyu Jia / Zehui Duan / Ping Huang / Sven T Stripp / Christopher K Barlow / Matthew Belousoff / Hannah S Shafaat / Gregory M Cook / Ralf B Schittenhelm / Kylie A Vincent / Syma Khalid / Gustav Berggren / Chris Greening /
Abstract: Diverse aerobic bacteria use atmospheric H as an energy source for growth and survival. This globally significant process regulates the composition of the atmosphere, enhances soil biodiversity and ...Diverse aerobic bacteria use atmospheric H as an energy source for growth and survival. This globally significant process regulates the composition of the atmosphere, enhances soil biodiversity and drives primary production in extreme environments. Atmospheric H oxidation is attributed to uncharacterized members of the [NiFe] hydrogenase superfamily. However, it remains unresolved how these enzymes overcome the extraordinary catalytic challenge of oxidizing picomolar levels of H amid ambient levels of the catalytic poison O and how the derived electrons are transferred to the respiratory chain. Here we determined the cryo-electron microscopy structure of the Mycobacterium smegmatis hydrogenase Huc and investigated its mechanism. Huc is a highly efficient oxygen-insensitive enzyme that couples oxidation of atmospheric H to the hydrogenation of the respiratory electron carrier menaquinone. Huc uses narrow hydrophobic gas channels to selectively bind atmospheric H at the expense of O, and 3 [3Fe-4S] clusters modulate the properties of the enzyme so that atmospheric H oxidation is energetically feasible. The Huc catalytic subunits form an octameric 833 kDa complex around a membrane-associated stalk, which transports and reduces menaquinone 94 Å from the membrane. These findings provide a mechanistic basis for the biogeochemically and ecologically important process of atmospheric H oxidation, uncover a mode of energy coupling dependent on long-range quinone transport, and pave the way for the development of catalysts that oxidize H in ambient air.
History
DepositionJul 20, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 4, 2023Provider: repository / Type: Initial release
Revision 1.1Jan 25, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.year
Revision 1.2Mar 8, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.title / _citation.year
Revision 1.3Mar 22, 2023Group: Database references / Category: citation / Item: _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.4Mar 29, 2023Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Hydrogenase-2, large subunit
B: Hydrogenase-2, small subunit
C: Hydrogenase-2, large subunit
D: Hydrogenase-2, small subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)187,16720
Polymers184,5784
Non-polymers2,58916
Water10,737596
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Hydrogenase-2, ... , 2 types, 4 molecules ACBD

#1: Protein Hydrogenase-2, large subunit /


Mass: 57217.078 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mycolicibacterium smegmatis (bacteria) / Strain: ATCC 700084 / mc(2)155 / References: UniProt: A0QUM7, hydrogenase (acceptor)
#2: Protein Hydrogenase-2, small subunit / / NADH ubiquinone oxidoreductase 20 kDa subunit


Mass: 35071.988 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mycolicibacterium smegmatis (bacteria) / Strain: ATCC 700084 / mc(2)155 / References: UniProt: I7G634, hydrogenase (acceptor)

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Non-polymers , 7 types, 612 molecules

#3: Chemical ChemComp-O / OXYGEN ATOM / Oxygen


Mass: 15.999 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: O / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical ChemComp-FCO / CARBONMONOXIDE-(DICYANO) IRON


Mass: 135.890 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C3FeN2O / Feature type: SUBJECT OF INVESTIGATION
#6: Chemical ChemComp-3NI / NICKEL (III) ION / Nickel


Mass: 58.693 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Formula: Ni / Feature type: SUBJECT OF INVESTIGATION
#7: Chemical ChemComp-VK3 / MENADIONE / VITAMIN K3 / 2-METHYL-1,4-NAPHTHALENEDIONE / Menadione


Mass: 172.180 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C11H8O2 / Feature type: SUBJECT OF INVESTIGATION
#8: Chemical
ChemComp-F3S / FE3-S4 CLUSTER / Iron–sulfur cluster


Mass: 295.795 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Fe3S4 / Feature type: SUBJECT OF INVESTIGATION
#9: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 596 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY

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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: Complex of the type 2 [NiFe]-hydrogenase Huc from Mycobacterium smegmatis
Type: COMPLEX / Entity ID: #1-#2 / Source: NATURAL
Molecular weightValue: 0.833 MDa / Experimental value: NO
Source (natural)Organism: Mycolicibacterium smegmatis (bacteria) / Strain: MC2 155
Buffer solutionpH: 7.9 / Details: pH 7.9
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMTris1
2200 mMSodium chlorideNaClSodium chloride1
SpecimenConc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid type: Quantifoil
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1500 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm
Specimen holderCryogen: HELIUM / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 4 sec. / Electron dose: 60.4 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of real images: 9868

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM software
IDNameVersionCategory
4CTFFIND4.1CTF correction
7UCSF ChimeraX1.3model fitting
8Coot0.92model fitting
10PHENIX1.2model refinement
11Coot0.92model refinement
12RELION3.1.2initial Euler assignment
13cryoSPARC3.3.2final Euler assignment
14cryoSPARC3.3.2classification
15cryoSPARC3.3.23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 3800000
3D reconstructionResolution: 1.52 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 153359 / Symmetry type: POINT
Atomic model buildingProtocol: BACKBONE TRACE / Space: REAL / Target criteria: Correlation coefficient
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00513452
ELECTRON MICROSCOPYf_angle_d0.59618332
ELECTRON MICROSCOPYf_dihedral_angle_d5.5911812
ELECTRON MICROSCOPYf_chiral_restr0.0421938
ELECTRON MICROSCOPYf_plane_restr0.0042396

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