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- EMDB-8193: Cryo-EM structure of isocitrate dehydrogenase (IDH1) in complex w... -

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Basic information

Entry
Database: EMDB / ID: EMD-8193
TitleCryo-EM structure of isocitrate dehydrogenase (IDH1) in complex with ML309 inhibitor
Map dataIsocitrate dehydrogenase (IDH1) in complex with ML309 inhibitor
Sample
  • Complex: Isocitrate dehydrogenase R132C mutant in complex with ML309
    • Protein or peptide: Isocitrate dehydrogenase [NADP] cytoplasmic
  • Ligand: NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE
Keywordsisocitrate dehydrogenase / small metabolic complex / small molecule inhibitor / OXIDOREDUCTASE
Function / homology
Function and homology information


Abnormal conversion of 2-oxoglutarate to 2-hydroxyglutarate / NADPH regeneration / regulation of phospholipid catabolic process / regulation of phospholipid biosynthetic process / NFE2L2 regulating TCA cycle genes / isocitrate metabolic process / isocitrate dehydrogenase (NADP+) / isocitrate dehydrogenase (NADP+) activity / 2-oxoglutarate metabolic process / NADP metabolic process ...Abnormal conversion of 2-oxoglutarate to 2-hydroxyglutarate / NADPH regeneration / regulation of phospholipid catabolic process / regulation of phospholipid biosynthetic process / NFE2L2 regulating TCA cycle genes / isocitrate metabolic process / isocitrate dehydrogenase (NADP+) / isocitrate dehydrogenase (NADP+) activity / 2-oxoglutarate metabolic process / NADP metabolic process / glyoxylate cycle / response to steroid hormone / female gonad development / peroxisomal matrix / tricarboxylic acid cycle / glutathione metabolic process / Peroxisomal protein import / NAD binding / peroxisome / tertiary granule lumen / NADP binding / secretory granule lumen / ficolin-1-rich granule lumen / response to oxidative stress / cadherin binding / Neutrophil degranulation / magnesium ion binding / protein homodimerization activity / mitochondrion / extracellular exosome / extracellular region / identical protein binding / cytosol / cytoplasm
Similarity search - Function
Isocitrate dehydrogenase NADP-dependent / Isocitrate/isopropylmalate dehydrogenase, conserved site / Isocitrate and isopropylmalate dehydrogenases signature. / Isopropylmalate dehydrogenase-like domain / Isocitrate/isopropylmalate dehydrogenase / Isocitrate/isopropylmalate dehydrogenase
Similarity search - Domain/homology
Isocitrate dehydrogenase [NADP] cytoplasmic
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsMerk A / Bartesaghi A
CitationJournal: Cell / Year: 2016
Title: Breaking Cryo-EM Resolution Barriers to Facilitate Drug Discovery.
Authors: Alan Merk / Alberto Bartesaghi / Soojay Banerjee / Veronica Falconieri / Prashant Rao / Mindy I Davis / Rajan Pragani / Matthew B Boxer / Lesley A Earl / Jacqueline L S Milne / Sriram Subramaniam /
Abstract: Recent advances in single-particle cryoelecton microscopy (cryo-EM) are enabling generation of numerous near-atomic resolution structures for well-ordered protein complexes with sizes ≥ ∼200 kDa. ...Recent advances in single-particle cryoelecton microscopy (cryo-EM) are enabling generation of numerous near-atomic resolution structures for well-ordered protein complexes with sizes ≥ ∼200 kDa. Whether cryo-EM methods are equally useful for high-resolution structural analysis of smaller, dynamic protein complexes such as those involved in cellular metabolism remains an important question. Here, we present 3.8 Å resolution cryo-EM structures of the cancer target isocitrate dehydrogenase (93 kDa) and identify the nature of conformational changes induced by binding of the allosteric small-molecule inhibitor ML309. We also report 2.8-Å- and 1.8-Å-resolution structures of lactate dehydrogenase (145 kDa) and glutamate dehydrogenase (334 kDa), respectively. With these results, two perceived barriers in single-particle cryo-EM are overcome: (1) crossing 2 Å resolution and (2) obtaining structures of proteins with sizes < 100 kDa, demonstrating that cryo-EM can be used to investigate a broad spectrum of drug-target interactions and dynamic conformational states.
History
DepositionMay 31, 2016-
Header (metadata) releaseJun 8, 2016-
Map releaseJun 8, 2016-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.00846
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.00846
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-5k11
  • Surface level: 0.00846
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_8193.png

Downloads & links

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Map

FileDownload / File: emd_8193.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationIsocitrate dehydrogenase (IDH1) in complex with ML309 inhibitor
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.5 Å/pix.
x 200 pix.
= 99. Å		0.5 Å/pix.
x 200 pix.
= 99. Å		0.5 Å/pix.
x 200 pix.
= 99. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.495 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.00846 / Movie #1: 0.00846
Minimum - Maximum-0.0077934233 - 0.021091789
Average (Standard dev.)0.000120903394 (±0.0025294467)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 99.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.4950.4950.495
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z99.00099.00099.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.0080.0210.000

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Supplemental data

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Additional map: Map sharpened using a B-factor of -180

Fileemd_8193_additional_1.map
AnnotationMap sharpened using a B-factor of -180
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Reconstruction obtained without imposing symmetry

Fileemd_8193_additional_2.map
AnnotationReconstruction obtained without imposing symmetry
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Isocitrate dehydrogenase R132C mutant in complex with ML309

EntireName: Isocitrate dehydrogenase R132C mutant in complex with ML309
Components
  • Complex: Isocitrate dehydrogenase R132C mutant in complex with ML309
    • Protein or peptide: Isocitrate dehydrogenase [NADP] cytoplasmic
  • Ligand: NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE

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Supramolecule #1: Isocitrate dehydrogenase R132C mutant in complex with ML309

SupramoleculeName: Isocitrate dehydrogenase R132C mutant in complex with ML309
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 93 KDa

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Macromolecule #1: Isocitrate dehydrogenase [NADP] cytoplasmic

MacromoleculeName: Isocitrate dehydrogenase [NADP] cytoplasmic / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: isocitrate dehydrogenase (NADP+)
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 46.334742 KDa
Recombinant expressionOrganism: unidentified baculovirus
SequenceString: KKISGGSVVE MQGDEMTRII WELIKEKLIF PYVELDLHSY DLGIENRDAT NDQVTKDAAE AIKKHNVGVK CATITPDEKR VEEFKLKQM WKSPNGTIRN ILGGTVFREA IICKNIPRLV SGWVKPIIIG CHAYGDQYRA TDFVVPGPGK VEITYTPSDG T QKVTYLVH ...String:
KKISGGSVVE MQGDEMTRII WELIKEKLIF PYVELDLHSY DLGIENRDAT NDQVTKDAAE AIKKHNVGVK CATITPDEKR VEEFKLKQM WKSPNGTIRN ILGGTVFREA IICKNIPRLV SGWVKPIIIG CHAYGDQYRA TDFVVPGPGK VEITYTPSDG T QKVTYLVH NFEEGGGVAM GMYNQDKSIE DFAHSSFQMA LSKGWPLYLS TKNTILKKYD GRFKDIFQEI YDKQYKSQFE AQ KIWYEHR LIDDMVAQAM KSEGGFIWAC KNYDGDVQSD SVAQGYGSLG MMTSVLVCPD GKTVEAEAAH GTVTRHYRMY QKG QETSTN PIASIFAWTR GLAHRAKLDN NKELAFFANA LEEVSIETIE AGFMTKDLAA CIKGLPNVQR SDYLNTFEFM DKLG ENLKI KLAQAK

UniProtKB: Isocitrate dehydrogenase [NADP] cytoplasmic

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Macromolecule #2: NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE

MacromoleculeName: NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE
type: ligand / ID: 2 / Number of copies: 2 / Formula: NDP
Molecular weightTheoretical: 745.421 Da
Chemical component information

ChemComp-NDP:
NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE

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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

Concentration2.8 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
20.0 mMC4H12NO3ClTris-HCl
150.0 mMNaClsodium chloride
10.0 mMMgCl2magnesium chloride
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE / Instrument: LEICA EM GP / Details: Plunged into liquid ethane (LEICA EM GP).

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Electron microscopy

MicroscopeFEI TITAN KRIOS
TemperatureMin: 79.6 K / Max: 79.8 K
Specialist opticsEnergy filter - Name: GIF Quantum / Energy filter - Lower energy threshold: 0 eV / Energy filter - Upper energy threshold: 20 eV
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 0-29 / Number real images: 820 / Average exposure time: 0.2 sec. / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 101000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.6 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 270000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 232343
Startup modelType of model: OTHER
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: FREALIGN (ver. 9.10)
Details: The primary map for this entry corresponds to the uncorrected reconstruction. A version sharpened using a B-factor of -180 is provided as additional volume data. The reconstruction, obtained ...Details: The primary map for this entry corresponds to the uncorrected reconstruction. A version sharpened using a B-factor of -180 is provided as additional volume data. The reconstruction, obtained without imposing symmetry, is also provided with this entry as additional volume data.
Number images used: 46483
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: FREALIGN (ver. 9.10)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: FREALIGN (ver. 9.10)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-5k11:
Cryo-EM structure of isocitrate dehydrogenase (IDH1) in inhibitor-bound state

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