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- EMDB-3733: Minor class in a preparation of mitochondrial complex I set in th... -

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Basic information

Entry
Database: EMDB / ID: EMD-3733
TitleMinor class in a preparation of mitochondrial complex I set in the deactive state
Map dataA minor class found in a preparation of mitochondrial complex I set in the deactive state.
Sample
  • Complex: Minor class found in a preparation of mitochondrial complex I set in the deactive state
Biological speciesBos taurus (cattle)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.5 Å
AuthorsBlaza JN / Vinothkumar KR / Hirst J
CitationJournal: Structure / Year: 2018
Title: Structure of the Deactive State of Mammalian Respiratory Complex I.
Authors: James N Blaza / Kutti R Vinothkumar / Judy Hirst /
Abstract: Complex I (NADH:ubiquinone oxidoreductase) is central to energy metabolism in mammalian mitochondria. It couples NADH oxidation by ubiquinone to proton transport across the energy-conserving inner ...Complex I (NADH:ubiquinone oxidoreductase) is central to energy metabolism in mammalian mitochondria. It couples NADH oxidation by ubiquinone to proton transport across the energy-conserving inner membrane, catalyzing respiration and driving ATP synthesis. In the absence of substrates, active complex I gradually enters a pronounced resting or deactive state. The active-deactive transition occurs during ischemia and is crucial for controlling how respiration recovers upon reperfusion. Here, we set a highly active preparation of Bos taurus complex I into the biochemically defined deactive state, and used single-particle electron cryomicroscopy to determine its structure to 4.1 Å resolution. We show that the deactive state arises when critical structural elements that form the ubiquinone-binding site become disordered, and we propose reactivation is induced when substrate binding to the NADH-reduced enzyme templates their reordering. Our structure both rationalizes biochemical data on the deactive state and offers new insights into its physiological and cellular roles.
History
DepositionMay 23, 2017-
Header (metadata) releaseMay 31, 2017-
Map releaseJan 17, 2018-
UpdateMar 21, 2018-
Current statusMar 21, 2018Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.12
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.12
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3733.png

Downloads & links

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Map

FileDownload / File: emd_3733.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationA minor class found in a preparation of mitochondrial complex I set in the deactive state.
Voxel sizeX=Y=Z: 1.38 Å
Density
Contour LevelBy AUTHOR: 0.12 / Movie #1: 0.12
Minimum - Maximum-0.12779298 - 0.5350517
Average (Standard dev.)0.0004159258 (±0.016465593)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 496.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.381.381.38
M x/y/z360360360
origin x/y/z0.0000.0000.000
length x/y/z496.800496.800496.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS360360360
D min/max/mean-0.1280.5350.000

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Supplemental data

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Sample components

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Entire : Minor class found in a preparation of mitochondrial complex I set...

EntireName: Minor class found in a preparation of mitochondrial complex I set in the deactive state
Components
  • Complex: Minor class found in a preparation of mitochondrial complex I set in the deactive state

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Supramolecule #1: Minor class found in a preparation of mitochondrial complex I set...

SupramoleculeName: Minor class found in a preparation of mitochondrial complex I set in the deactive state
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#44
Source (natural)Organism: Bos taurus (cattle)
Molecular weightExperimental: 1.0 MDa

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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

Concentration4 mg/mL
BufferpH: 7.55
Component:
ConcentrationNameFormula
20.0 mMTris-Cl
150.0 mMNaClSodium chloride
0.04 %Cymal-7
GridModel: Quantifoil UltrAuFoil R0.6/1 / Material: GOLD / Details: The grid was PEGylated before use
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
DetailsMonodisperse bovine mitochondrial complex I isolated in the detergent cymal-7

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated magnification: 101499 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.1 µm / Nominal defocus min: 1.3 µm / Nominal magnification: 59000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Digitization - Sampling interval: 14.0 µm / Digitization - Frames/image: 1-20 / Number grids imaged: 1 / Number real images: 2954 / Average exposure time: 2.5 sec. / Average electron dose: 80.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 148488
CTF correctionSoftware - Name: RELION / Details: CTF parameters were applied internally in Relion
Startup modelType of model: EMDB MAP
EMDB ID:

2676


Details: Filtred to 60 A
Initial angle assignmentType: NOT APPLICABLE
Final 3D classificationNumber classes: 3 / Software - Name: RELION (ver. 1.4) / Details: Running with 6 classes gave very similar results.
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 1.4)
Details: Relion uses projection matching in a ML statistical framework
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 7.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.4) / Number images used: 12048
DetailsImages were normalised in Relion
FSC plot (resolution estimation)

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