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- PDB-7nz1: Respiratory complex I from Escherichia coli - focused refinement ... -

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Basic information

Entry
Database: PDB / ID: 7nz1
TitleRespiratory complex I from Escherichia coli - focused refinement of cytoplasmic arm
Components(NADH-quinone oxidoreductase subunit ...NADH dehydrogenase (quinone)) x 6
KeywordsELECTRON TRANSPORT / NADH:ubiquinone reductase (H+-translocating) / oxidative phosphorylation
Function / homology
Function and homology information


NADH dehydrogenase complex / plasma membrane respiratory chain complex I / Translocases; Catalysing the translocation of protons; Linked to oxidoreductase reactions / NADH:ubiquinone reductase (non-electrogenic) activity / molybdopterin cofactor binding / respiratory chain complex I / cellular respiration / NADH dehydrogenase activity / NADH dehydrogenase (ubiquinone) activity / quinone binding ...NADH dehydrogenase complex / plasma membrane respiratory chain complex I / Translocases; Catalysing the translocation of protons; Linked to oxidoreductase reactions / NADH:ubiquinone reductase (non-electrogenic) activity / molybdopterin cofactor binding / respiratory chain complex I / cellular respiration / NADH dehydrogenase activity / NADH dehydrogenase (ubiquinone) activity / quinone binding / ATP synthesis coupled electron transport / aerobic respiration / respiratory electron transport chain / transmembrane transport / 2 iron, 2 sulfur cluster binding / NAD binding / FMN binding / 4 iron, 4 sulfur cluster binding / iron ion binding / membrane / metal ion binding / plasma membrane / cytoplasm
Similarity search - Function
NADH dehydrogenase, subunit CD / Molybdopterin oxidoreductase Fe4S4 domain / Molybdopterin oxidoreductase Fe4S4 domain / Molybdopterin dinucleotide-binding domain / Molydopterin dinucleotide binding domain / NADH-quinone oxidoreductase, chain I / NAD(P)H-quinone oxidoreductase subunit D/H / NADH:ubiquinone oxidoreductase, 49kDa subunit, conserved site / Respiratory chain NADH dehydrogenase 49 Kd subunit signature. / NADH-quinone oxidoreductase, subunit D ...NADH dehydrogenase, subunit CD / Molybdopterin oxidoreductase Fe4S4 domain / Molybdopterin oxidoreductase Fe4S4 domain / Molybdopterin dinucleotide-binding domain / Molydopterin dinucleotide binding domain / NADH-quinone oxidoreductase, chain I / NAD(P)H-quinone oxidoreductase subunit D/H / NADH:ubiquinone oxidoreductase, 49kDa subunit, conserved site / Respiratory chain NADH dehydrogenase 49 Kd subunit signature. / NADH-quinone oxidoreductase, subunit D / Respiratory-chain NADH dehydrogenase, 49 Kd subunit / NADH:ubiquinone oxidoreductase, subunit G / Respiratory-chain NADH dehydrogenase 75 Kd subunit signature 3. / Respiratory-chain NADH dehydrogenase 75 Kd subunit signature 2. / NADH dehydrogenase, subunit C / NADH ubiquinone oxidoreductase, F subunit / NADH:ubiquinone oxidoreductase, 30kDa subunit, conserved site / Respiratory chain NADH dehydrogenase 30 Kd subunit signature. / 2Fe-2S iron-sulfur cluster binding domain / Respiratory-chain NADH dehydrogenase 75 Kd subunit signature 1. / NADH:ubiquinone oxidoreductase, 75kDa subunit, conserved site / NADH:ubiquinone oxidoreductase, 30kDa subunit / NADH:ubiquinone oxidoreductase, 30kDa subunit superfamily / Respiratory-chain NADH dehydrogenase, 30 Kd subunit / NADH-ubiquinone oxidoreductase-G iron-sulfur binding region / NADH-ubiquinone oxidoreductase-G iron-sulfur binding region / Respiratory-chain NADH dehydrogenase 51 Kd subunit signature 1. / Respiratory-chain NADH dehydrogenase 24 Kd subunit signature. / Aspartate decarboxylase-like domain superfamily / NuoE domain / NADH-quinone oxidoreductase subunit E-like / Thioredoxin-like [2Fe-2S] ferredoxin / NADH-quinone oxidoreductase subunit E, N-terminal / NADH:ubiquinone oxidoreductase, subunit G, iron-sulphur binding / His(Cys)3-ligated-type [4Fe-4S] domain profile. / NADH:ubiquinone oxidoreductase, 51kDa subunit, conserved site / Respiratory-chain NADH dehydrogenase 51 Kd subunit signature 2. / NADH-ubiquinone oxidoreductase 51kDa subunit, FMN-binding domain / NADH-ubiquinone oxidoreductase 51kDa subunit, iron-sulphur binding domain / NADH-ubiquinone oxidoreductase 51kDa subunit, iron-sulphur binding domain superfamily / NADH-ubiquinone oxidoreductase 51kDa subunit, FMN-binding domain superfamily / Respiratory-chain NADH dehydrogenase 51 Kd subunit / NADH-ubiquinone oxidoreductase-F iron-sulfur binding region / NADH-ubiquinone oxidoreductase-F iron-sulfur binding region / Molybdopterin oxidoreductase, 4Fe-4S domain / Prokaryotic molybdopterin oxidoreductases 4Fe-4S domain profile. / Molybdopterin oxidoreductase / Molybdopterin oxidoreductase / [NiFe]-hydrogenase, large subunit / 4Fe-4S dicluster domain / 2Fe-2S ferredoxin-type iron-sulfur binding domain profile. / 2Fe-2S ferredoxin-type iron-sulfur binding domain / 2Fe-2S ferredoxin-like superfamily / 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 / Thioredoxin-like superfamily
Similarity search - Domain/homology
FE2/S2 (INORGANIC) CLUSTER / FLAVIN MONONUCLEOTIDE / IRON/SULFUR CLUSTER / NADH-quinone oxidoreductase subunit E / NADH-quinone oxidoreductase subunit I / NADH-quinone oxidoreductase subunit F / NADH-quinone oxidoreductase subunit C/D / NADH-quinone oxidoreductase subunit G
Similarity search - Component
Biological speciesEscherichia coli B (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.1 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 23, 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
B: NADH-quinone oxidoreductase subunit B
D: NADH-quinone oxidoreductase subunit C/D
E: NADH-quinone oxidoreductase subunit E
F: NADH-quinone oxidoreductase subunit F
G: NADH-quinone oxidoreductase subunit G
I: NADH-quinone oxidoreductase subunit I
hetero molecules


Theoretical massNumber of molelcules
Total (without water)285,71017
Polymers282,4006
Non-polymers3,31011
Water21,0781170
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area31730 Å2
ΔGint-315 kcal/mol
Surface area77950 Å2
MethodPISA

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Components

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NADH-quinone oxidoreductase subunit ... , 6 types, 6 molecules BDEFGI

#1: Protein NADH-quinone oxidoreductase subunit B / NADH dehydrogenase (quinone)


Mass: 25097.809 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / References: NADH:ubiquinone reductase (H+-translocating)
#2: Protein NADH-quinone oxidoreductase subunit C/D / NADH dehydrogenase (quinone) / NADH dehydrogenase I subunit C/D / NDH-1 subunit C/D / NUO3/NUO4


Mass: 68321.945 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: BL21(AI)
References: UniProt: P33599, NADH:ubiquinone reductase (H+-translocating)
#3: Protein NADH-quinone oxidoreductase subunit E / NADH dehydrogenase (quinone)


Mass: 18630.049 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: P0AFD1, NADH:ubiquinone reductase (H+-translocating)
#4: Protein NADH-quinone oxidoreductase subunit F / NADH dehydrogenase (quinone)


Mass: 49368.332 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: P31979, NADH:ubiquinone reductase (H+-translocating)
#5: Protein NADH-quinone oxidoreductase subunit G / NADH dehydrogenase (quinone)


Mass: 100419.211 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: P33602, NADH:ubiquinone reductase (H+-translocating)
#6: Protein NADH-quinone oxidoreductase subunit I / NADH dehydrogenase (quinone)


Mass: 20562.771 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria)
References: UniProt: P0AFD6, NADH:ubiquinone reductase (H+-translocating)

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Non-polymers , 5 types, 1181 molecules

#7: Chemical
ChemComp-SF4 / IRON/SULFUR CLUSTER / Iron–sulfur cluster


Mass: 351.640 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: Fe4S4
#8: Chemical ChemComp-FES / FE2/S2 (INORGANIC) CLUSTER / Iron–sulfur cluster


Mass: 175.820 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Fe2S2
#9: Chemical ChemComp-FMN / FLAVIN MONONUCLEOTIDE / RIBOFLAVIN MONOPHOSPHATE / Flavin mononucleotide


Mass: 456.344 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C17H21N4O9P
#10: Chemical ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Ca
#11: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 1170 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestN

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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 cytoplasmic arm
Type: COMPLEX / Entity ID: #1-#6 / Source: NATURAL
Molecular weightValue: 0.28 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: 2.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 286333 / Symmetry type: POINT
Atomic model buildingB value: 30 / Protocol: AB INITIO MODEL / Space: REAL / Target criteria: Correlation coefficient
Atomic model buildingPDB-ID: 4HEA

4hea

RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 30.33 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.005819062
ELECTRON MICROSCOPYf_angle_d0.936925882
ELECTRON MICROSCOPYf_chiral_restr0.05822787
ELECTRON MICROSCOPYf_plane_restr0.00773381
ELECTRON MICROSCOPYf_dihedral_angle_d15.9887034

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