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- EMDB-10584: PKM2 in complex with Compound 10 -

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Basic information

Entry
Database: EMDB / ID: EMD-10584
TitlePKM2 in complex with Compound 10
Map dataNone
Sample
  • Complex: Pyruvate Kinase M2
    • Protein or peptide: Pyruvate kinase PKM
  • Ligand: 1-propan-2-yl-3-pyridin-4-yl-urea
  • Ligand: 1,6-di-O-phosphono-beta-D-fructofuranose
  • Ligand: water
Function / homology
Function and homology information


programmed cell death / pyruvate kinase / pyruvate kinase activity / positive regulation of cytoplasmic translation / histone H3T11 kinase activity / canonical glycolysis / Glycolysis / positive regulation of sprouting angiogenesis / potassium ion binding / rough endoplasmic reticulum ...programmed cell death / pyruvate kinase / pyruvate kinase activity / positive regulation of cytoplasmic translation / histone H3T11 kinase activity / canonical glycolysis / Glycolysis / positive regulation of sprouting angiogenesis / potassium ion binding / rough endoplasmic reticulum / glycolytic process / non-specific protein-tyrosine kinase / cilium / cellular response to insulin stimulus / extracellular vesicle / MHC class II protein complex binding / protein tyrosine kinase activity / collagen-containing extracellular matrix / secretory granule lumen / vesicle / ficolin-1-rich granule lumen / transcription coactivator activity / non-specific serine/threonine protein kinase / cadherin binding / phosphorylation / mRNA binding / Neutrophil degranulation / magnesium ion binding / protein homodimerization activity / positive regulation of transcription by RNA polymerase II / mitochondrion / RNA binding / extracellular exosome / extracellular region / ATP binding / nucleus / cytosol / cytoplasm
Similarity search - Function
Pyruvate kinase, active site / Pyruvate kinase active site signature. / Pyruvate kinase / Pyruvate kinase, barrel / Pyruvate kinase, insert domain superfamily / Pyruvate kinase, barrel domain / Pyruvate kinase, C-terminal / Pyruvate kinase, C-terminal domain superfamily / Pyruvate kinase, alpha/beta domain / Pyruvate kinase-like, insert domain superfamily ...Pyruvate kinase, active site / Pyruvate kinase active site signature. / Pyruvate kinase / Pyruvate kinase, barrel / Pyruvate kinase, insert domain superfamily / Pyruvate kinase, barrel domain / Pyruvate kinase, C-terminal / Pyruvate kinase, C-terminal domain superfamily / Pyruvate kinase, alpha/beta domain / Pyruvate kinase-like, insert domain superfamily / Pyruvate kinase-like domain superfamily / Pyruvate/Phosphoenolpyruvate kinase-like domain superfamily
Similarity search - Domain/homology
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsSaur M / Hartshorn MJ / Dong J / Reeks J / Bunkoczi G / Jhoti H / Williams PA
CitationJournal: Drug Discov Today / Year: 2020
Title: Fragment-based drug discovery using cryo-EM.
Authors: Michael Saur / Michael J Hartshorn / Jing Dong / Judith Reeks / Gabor Bunkoczi / Harren Jhoti / Pamela A Williams /
Abstract: Recent advances in electron cryo-microscopy (cryo-EM) structure determination have pushed the resolutions obtainable by the method into the range widely considered to be of utility for drug discovery. ...Recent advances in electron cryo-microscopy (cryo-EM) structure determination have pushed the resolutions obtainable by the method into the range widely considered to be of utility for drug discovery. Here, we review the use of cryo-EM in fragment-based drug discovery (FBDD) based on in-house method development. We demonstrate not only that cryo-EM can reveal details of the molecular interactions between fragments and a protein, but also that the current reproducibility, quality, and throughput are compatible with FBDD. We exemplify this using the test system β-galactosidase (Bgal) and the oncology target pyruvate kinase 2 (PKM2).
History
DepositionDec 30, 2019-
Header (metadata) releaseJan 15, 2020-
Map releaseJan 15, 2020-
UpdateJan 27, 2021-
Current statusJan 27, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.05
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6ttq
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10584.png

Downloads & links

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Map

FileDownload / File: emd_10584.map.gz / Format: CCP4 / Size: 39.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNone
Voxel sizeX=Y=Z: 0.5 Å
Density
Contour LevelBy AUTHOR: 0.05 / Movie #1: 0.05
Minimum - Maximum-0.2144623 - 0.37817344
Average (Standard dev.)0.00357944 (±0.026130913)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions223175263
Spacing175223263
CellA: 87.5 Å / B: 111.5 Å / C: 131.5 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.50.50.5
M x/y/z175223263
origin x/y/z0.0000.0000.000
length x/y/z87.500111.500131.500
α/β/γ90.00090.00090.000
start NX/NY/NZ1081340
NX/NY/NZ13584349
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS175223263
D min/max/mean-0.2140.3780.004

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Supplemental data

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Additional map: Relion post-process unmasked map from halfmaps which have...

Fileemd_10584_additional_1.map
AnnotationRelion post-process unmasked map from halfmaps which have not been resampled after auto-refine.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: None

Fileemd_10584_additional_2.map
AnnotationNone
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Relion auto-refine halfmap 1

Fileemd_10584_half_map_1.map
AnnotationRelion auto-refine halfmap 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Relion auto-refine halfmap 2

Fileemd_10584_half_map_2.map
AnnotationRelion auto-refine halfmap 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Pyruvate Kinase M2

EntireName: Pyruvate Kinase M2
Components
  • Complex: Pyruvate Kinase M2
    • Protein or peptide: Pyruvate kinase PKM
  • Ligand: 1-propan-2-yl-3-pyridin-4-yl-urea
  • Ligand: 1,6-di-O-phosphono-beta-D-fructofuranose
  • Ligand: water

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Supramolecule #1: Pyruvate Kinase M2

SupramoleculeName: Pyruvate Kinase M2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli)
Molecular weightTheoretical: 240 KDa

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Macromolecule #1: Pyruvate kinase PKM

MacromoleculeName: Pyruvate kinase PKM / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO / EC number: pyruvate kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 59.83182 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MGSSHHHHHH SSGLVPRGSK PHSEAGTAFI QTQQLHAAMA DTFLEHMCRL DIDSPPITAR NTGIICTIGP ASRSVETLKE MIKSGMNVA RLNFSHGTHE YHAETIKNVR TATESFASDP ILYRPVAVAL DTKGPEIRTG LIKGSGTAEV ELKKGATLKI T LDNAYMEK ...String:
MGSSHHHHHH SSGLVPRGSK PHSEAGTAFI QTQQLHAAMA DTFLEHMCRL DIDSPPITAR NTGIICTIGP ASRSVETLKE MIKSGMNVA RLNFSHGTHE YHAETIKNVR TATESFASDP ILYRPVAVAL DTKGPEIRTG LIKGSGTAEV ELKKGATLKI T LDNAYMEK CDENILWLDY KNICKVVEVG SKIYVDDGLI SLQVKQKGAD FLVTEVENGG SLGSKKGVNL PGAAVDLPAV SE KDIQDLK FGVEQDVDMV FASFIRKASD VHEVRKVLGE KGKNIKIISK IENHEGVRRF DEILEASDGI MVARGDLGIE IPA EKVFLA QKMMIGRCNR AGKPVICATQ MLESMIKKPR PTRAEGSDVA NAVLDGADCI MLSGETAKGD YPLEAVRMQH LIAR EAEAA IYHLQLFEEL RRLAPITSDP TEATAVGAVE ASFKCCSGAI IVLTKSGRSA HQVARYRPRA PIIAVTRNPQ TARQA HLYR GIFPVLCKDP VQEAWAEDVD LRVNFAMNVG KARGFFKKGD VVIVLTGWRP GSGFTNTMRV VPVP

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Macromolecule #2: 1-propan-2-yl-3-pyridin-4-yl-urea

MacromoleculeName: 1-propan-2-yl-3-pyridin-4-yl-urea / type: ligand / ID: 2 / Number of copies: 4 / Formula: NXH
Molecular weightTheoretical: 179.219 Da
Chemical component information

ChemComp-NXH:
1-propan-2-yl-3-pyridin-4-yl-urea

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Macromolecule #3: 1,6-di-O-phosphono-beta-D-fructofuranose

MacromoleculeName: 1,6-di-O-phosphono-beta-D-fructofuranose / type: ligand / ID: 3 / Number of copies: 4 / Formula: FBP
Molecular weightTheoretical: 340.116 Da
Chemical component information

ChemComp-FBP:
1,6-di-O-phosphono-beta-D-fructofuranose / Fructose 1,6-bisphosphate

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 28 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER / Water

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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

Concentration0.16 mg/mL
BufferpH: 8.2
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: GRAPHENE OXIDE / Support film - topology: CONTINUOUS
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 572 / Average exposure time: 59.98 sec. / Average electron dose: 65.4 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 267920
Startup modelType of model: OTHER / Details: in-house X-ray structure
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: D2 (2x2 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.04) / Number images used: 142864

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Atomic model buiding 1

RefinementSpace: RECIPROCAL / Protocol: OTHER / Overall B value: 75.429 / Target criteria: Maximum likelihood with phases
Output model

PDB-6ttq:
PKM2 in complex with Compound 10

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