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- PDB-7nyh: Respiratory complex I from Escherichia coli - focused refinement ... -

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Basic information

Entry
Database: PDB / ID: 7nyh
TitleRespiratory complex I from Escherichia coli - focused refinement of membrane arm
Components(NADH-quinone oxidoreductase subunit ...NADH dehydrogenase (quinone)) x 7
KeywordsELECTRON TRANSPORT / NADH:ubiquinone reductase (H+-translocating) / oxidative phosphorylation
Function / homology
Function and homology information


plasma membrane respiratory chain complex I / NADH dehydrogenase complex / Translocases; Catalysing the translocation of protons; Linked to oxidoreductase reactions / NADH:ubiquinone reductase (non-electrogenic) activity / respiratory chain complex I / ubiquinone binding / oxidoreductase activity, acting on NAD(P)H, quinone or similar compound as acceptor / electron transport coupled proton transport / NADH dehydrogenase (ubiquinone) activity / quinone binding ...plasma membrane respiratory chain complex I / NADH dehydrogenase complex / Translocases; Catalysing the translocation of protons; Linked to oxidoreductase reactions / NADH:ubiquinone reductase (non-electrogenic) activity / respiratory chain complex I / ubiquinone binding / oxidoreductase activity, acting on NAD(P)H, quinone or similar compound as acceptor / electron transport coupled proton transport / NADH dehydrogenase (ubiquinone) activity / quinone binding / ATP synthesis coupled electron transport / aerobic respiration / respiratory electron transport chain / membrane => GO:0016020 / membrane / plasma membrane
Similarity search - Function
NAD(P)H-quinone oxidoreductase subunit 3, bacterial/plastid / NAD(P)H-quinone oxidoreductase, subunit N/subunit 2 / NADH-ubiquinone/plastoquinone oxidoreductase chain 6, subunit NuoJ / NADH-plastoquinone oxidoreductase, chain 5 subgroup / NADH-quinone oxidoreductase, chain M/4 / NADH-ubiquinone oxidoreductase chain 4L/K / NADH:ubiquinone/plastoquinone oxidoreductase, chain 6 / NADH-ubiquinone/plastoquinone oxidoreductase chain 6 / NADH-Ubiquinone oxidoreductase (complex I), chain 5 N-terminal / NADH-quinone oxidoreductase, chain 5-like ...NAD(P)H-quinone oxidoreductase subunit 3, bacterial/plastid / NAD(P)H-quinone oxidoreductase, subunit N/subunit 2 / NADH-ubiquinone/plastoquinone oxidoreductase chain 6, subunit NuoJ / NADH-plastoquinone oxidoreductase, chain 5 subgroup / NADH-quinone oxidoreductase, chain M/4 / NADH-ubiquinone oxidoreductase chain 4L/K / NADH:ubiquinone/plastoquinone oxidoreductase, chain 6 / NADH-ubiquinone/plastoquinone oxidoreductase chain 6 / NADH-Ubiquinone oxidoreductase (complex I), chain 5 N-terminal / NADH-quinone oxidoreductase, chain 5-like / NADH-Ubiquinone oxidoreductase (complex I), chain 5 N-terminus / NADH-ubiquinone oxidoreductase chain 4L/Mnh complex subunit C1-like / NADH-ubiquinone/plastoquinone oxidoreductase chain 4L / NADH:ubiquinone/plastoquinone oxidoreductase, chain 3 / NADH:ubiquinone oxidoreductase, subunit 3 superfamily / NADH-ubiquinone/plastoquinone oxidoreductase, chain 3 / NADH:ubiquinone oxidoreductase, subunit 1, conserved site / Respiratory-chain NADH dehydrogenase subunit 1 signature 1. / Respiratory-chain NADH dehydrogenase subunit 1 signature 2. / NADH:ubiquinone oxidoreductase, subunit 1/F420H2 oxidoreductase subunit H / NADH dehydrogenase / NADH:ubiquinone oxidoreductase / NADH:quinone oxidoreductase/Mrp antiporter, membrane subunit / Proton-conducting membrane transporter
Similarity search - Domain/homology
NADH-quinone oxidoreductase subunit L / NADH-quinone oxidoreductase subunit K / NADH-quinone oxidoreductase subunit A / NADH-quinone oxidoreductase subunit H / NADH-quinone oxidoreductase subunit J / NADH-quinone oxidoreductase subunit M / NADH-quinone oxidoreductase subunit N
Similarity search - Component
Biological speciesEscherichia coli B (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsKolata, P. / Efremov, R.G.
Funding support Belgium, 1items
OrganizationGrant numberCountry
European Research Council (ERC)00281ROEF Belgium
CitationJournal: Elife / Year: 2021
Title: Structure of respiratory complex I reconstituted into lipid nanodiscs reveals an uncoupled conformation.
Authors: Piotr Kolata / Rouslan G Efremov /
Abstract: Respiratory complex I is a multi-subunit membrane protein complex that reversibly couples NADH oxidation and ubiquinone reduction with proton translocation against transmembrane potential. Complex I ...Respiratory complex I is a multi-subunit membrane protein complex that reversibly couples NADH oxidation and ubiquinone reduction with proton translocation against transmembrane potential. Complex I from is among the best functionally characterized complexes, but its structure remains unknown, hindering further studies to understand the enzyme coupling mechanism. Here, we describe the single particle cryo-electron microscopy (cryo-EM) structure of the entire catalytically active complex I reconstituted into lipid nanodiscs. The structure of this mesophilic bacterial complex I displays highly dynamic connection between the peripheral and membrane domains. The peripheral domain assembly is stabilized by unique terminal extensions and an insertion loop. The membrane domain structure reveals novel dynamic features. Unusual conformation of the conserved interface between the peripheral and membrane domains suggests an uncoupled conformation of the complex. Considering constraints imposed by the structural data, we suggest a new simple hypothetical coupling mechanism for the molecular machine.
History
DepositionMar 22, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0Aug 18, 2021Provider: repository / Type: Initial release

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Structure visualization

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Assembly

Deposited unit
A: NADH-quinone oxidoreductase subunit A
H: NADH-quinone oxidoreductase subunit H
J: NADH-quinone oxidoreductase subunit J
K: NADH-quinone oxidoreductase subunit K
L: NADH-quinone oxidoreductase subunit L
M: NADH-quinone oxidoreductase subunit M
N: NADH-quinone oxidoreductase subunit N


Theoretical massNumber of molelcules
Total (without water)258,6047
Polymers258,6047
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area34290 Å2
ΔGint-372 kcal/mol
Surface area72290 Å2
MethodPISA

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Components

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NADH-quinone oxidoreductase subunit ... , 7 types, 7 molecules AHJKLMN

#1: Protein NADH-quinone oxidoreductase subunit A / NADH dehydrogenase (quinone) / NADH dehydrogenase I subunit A / NDH-1 subunit A / NUO1


Mass: 16474.283 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: P0AFC3, Translocases; Catalysing the translocation of protons; Linked to oxidoreductase reactions
#2: Protein NADH-quinone oxidoreductase subunit H / NADH dehydrogenase (quinone) / NADH dehydrogenase I subunit H / NDH-1 subunit H / NUO8


Mass: 36240.922 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: P0AFD4, Translocases; Catalysing the translocation of protons; Linked to oxidoreductase reactions
#3: Protein NADH-quinone oxidoreductase subunit J / NADH dehydrogenase (quinone) / NADH dehydrogenase I subunit J / NDH-1 subunit J / NUO10


Mass: 19889.551 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: P0AFE0, Translocases; Catalysing the translocation of protons; Linked to oxidoreductase reactions
#4: Protein NADH-quinone oxidoreductase subunit K / NADH dehydrogenase (quinone)


Mass: 10852.961 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: F4VE45, NADH:ubiquinone reductase (H+-translocating)
#5: Protein NADH-quinone oxidoreductase subunit L / NADH dehydrogenase (quinone) / NuoL_2 protein


Mass: 66513.633 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: A0A1V3W1N5, EC: 1.6.5.11, NADH:ubiquinone reductase (H+-translocating), NADH dehydrogenase (quinone)
#6: Protein NADH-quinone oxidoreductase subunit M / NADH dehydrogenase (quinone) / NADH dehydrogenase I subunit M / NDH-1 subunit M / NUO13


Mass: 56560.090 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: P0AFE8, Translocases; Catalysing the translocation of protons; Linked to oxidoreductase reactions
#7: Protein NADH-quinone oxidoreductase subunit N / NADH dehydrogenase (quinone) / NADH dehydrogenase I subunit N / NDH-1 subunit N / NUO14


Mass: 52072.672 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Plasmid details: cell membrane
References: UniProt: P0AFF0, Translocases; Catalysing the translocation of protons; Linked to oxidoreductase reactions

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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: Respiratory complex I from Escherichia coli - focused refinement of membrane arm
Type: COMPLEX / Entity ID: all / Source: NATURAL
Molecular weightValue: 0.26 MDa / Experimental value: NO
Source (natural)Organism: Escherichia coli B (bacteria) / Strain: BL21(AI) / Cellular location: cell membrane
Buffer solutionpH: 6.8
Details: The buffer was used for gel filtration of protein reconstituted in lipid nanodiscs
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMBis-TrisBis-tris methaneC8H19NO51
2200 mMSodium chlorideNaClSodium chloride1
32 mMCalcium chlorideCaCl21
SpecimenConc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1
VitrificationInstrument: GATAN CRYOPLUNGE 3 / Cryogen name: ETHANE / Humidity: 97 % / Chamber temperature: 296 K

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Electron microscopy imaging

MicroscopyModel: JEOL CRYO ARM 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 60000 X / Nominal defocus max: 2200 nm / Nominal defocus min: 900 nm / Cs: 2.55 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: JEOL CRYOSPECPORTER
Image recordingAverage exposure time: 3 sec. / Electron dose: 64.7 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of real images: 9122
EM imaging opticsEnergyfilter name: In-column Omega Filter / Energyfilter slit width: 20 eV

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
EM software
IDNameVersionCategory
1crYOLO1.4particle selection
4CTFFIND4.1CTF correction
7Coot0.9model fitting
9RELION3initial Euler assignment
10RELION3.1final Euler assignment
11RELION3.1classification
12RELION3.13D reconstruction
13PHENIX1.18.23D reconstruction
20PHENIX1.19.2model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1256734
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 37441 / Symmetry type: POINT
Atomic model buildingB value: 75 / Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: Correlation coefficient
Atomic model building
IDPDB-ID 3D fitting-ID
14HEA

4hea

1
23RKO

3rko

1
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 77.81 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.006216555
ELECTRON MICROSCOPYf_angle_d0.943122582
ELECTRON MICROSCOPYf_chiral_restr0.04892698
ELECTRON MICROSCOPYf_plane_restr0.0062778
ELECTRON MICROSCOPYf_dihedral_angle_d13.72345456

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