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- EMDB-26810: KDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent co... -

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Basic information

Entry
Database: EMDB / ID: EMD-26810
TitleKDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate
Map dataKDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate
Sample
  • Complex: KDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate
    • Protein or peptide: human histone H3
    • Protein or peptide: human histone H4
    • Protein or peptide: human histone H2A
    • Protein or peptide: human histone H2B
    • DNA: DNA 185-MER
    • DNA: DNA 185-MER
    • Protein or peptide: Lysine-specific demethylase 2B (KDM2B)
Biological speciesHomo sapiens (human) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsSpangler CJ / Skrajna A / Foley CA / Budziszewski GR / Azzam DN / James LI / Frye SV / McGinty RK
Funding support United States, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM133498 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA253730 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM139514 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA010305 United States
CitationJournal: Nat Chem Biol / Year: 2023
Title: Structural basis of paralog-specific KDM2A/B nucleosome recognition.
Authors: Cathy J Spangler / Aleksandra Skrajna / Caroline A Foley / Anh Nguyen / Gabrielle R Budziszewski / Dalal N Azzam / Eyla C Arteaga / Holly C Simmons / Charlotte B Smith / Nathaniel A Wesley / ...Authors: Cathy J Spangler / Aleksandra Skrajna / Caroline A Foley / Anh Nguyen / Gabrielle R Budziszewski / Dalal N Azzam / Eyla C Arteaga / Holly C Simmons / Charlotte B Smith / Nathaniel A Wesley / Emily M Wilkerson / Jeanne-Marie E McPherson / Dmitri Kireev / Lindsey I James / Stephen V Frye / Dennis Goldfarb / Robert K McGinty /
Abstract: The nucleosome acidic patch is a major interaction hub for chromatin, providing a platform for enzymes to dock and orient for nucleosome-targeted activities. To define the molecular basis of acidic ...The nucleosome acidic patch is a major interaction hub for chromatin, providing a platform for enzymes to dock and orient for nucleosome-targeted activities. To define the molecular basis of acidic patch recognition proteome wide, we performed an amino acid resolution acidic patch interactome screen. We discovered that the histone H3 lysine 36 (H3K36) demethylase KDM2A, but not its closely related paralog, KDM2B, requires the acidic patch for nucleosome binding. Despite fundamental roles in transcriptional repression in health and disease, the molecular mechanisms governing nucleosome substrate specificity of KDM2A/B, or any related JumonjiC (JmjC) domain lysine demethylase, remain unclear. We used a covalent conjugate between H3K36 and a demethylase inhibitor to solve cryogenic electron microscopy structures of KDM2A and KDM2B trapped in action on a nucleosome substrate. Our structures show that KDM2-nucleosome binding is paralog specific and facilitated by dynamic nucleosomal DNA unwrapping and histone charge shielding that mobilize the H3K36 sequence for demethylation.
History
DepositionApr 29, 2022-
Header (metadata) releaseFeb 22, 2023-
Map releaseFeb 22, 2023-
UpdateMay 17, 2023-
Current statusMay 17, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26810.png

Downloads & links

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Map

FileDownload / File: emd_26810.map.gz / Format: CCP4 / Size: 137.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationKDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.91 Å/pix.
x 330 pix.
= 300.3 Å		0.91 Å/pix.
x 330 pix.
= 300.3 Å		0.91 Å/pix.
x 330 pix.
= 300.3 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.91 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.018
Minimum - Maximum-0.03936551 - 0.07742096
Average (Standard dev.)0.0001535887 (±0.001865464)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions330330330
Spacing330330330
CellA=B=C: 300.30002 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: KDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate half...

Fileemd_26810_half_map_1.map
AnnotationKDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate half map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: KDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate half...

Fileemd_26810_half_map_2.map
AnnotationKDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate half map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : KDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent co...

EntireName: KDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate
Components
  • Complex: KDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate
    • Protein or peptide: human histone H3
    • Protein or peptide: human histone H4
    • Protein or peptide: human histone H2A
    • Protein or peptide: human histone H2B
    • DNA: DNA 185-MER
    • DNA: DNA 185-MER
    • Protein or peptide: Lysine-specific demethylase 2B (KDM2B)

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Supramolecule #1: KDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent co...

SupramoleculeName: KDM2B-nucleosome complex stabilized by H3K36C-UNC8015 covalent conjugate
type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: #1-#7
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 320 KDa

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Macromolecule #1: human histone H3

MacromoleculeName: human histone H3 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
ARTKQTARKS TGGKAPRKQL ATKAARKSAP ATGGVCKPHR YRPGTVALRE IRRYQKSTEL LIRKLPFQRL VREIAQDFKT DLRFQSSAVM ALQEASEAYL VGLFEDTNLA AIHAKRVTIM PKDIQLARRI RGERA

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Macromolecule #2: human histone H4

MacromoleculeName: human histone H4 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
SGRGKGGKGL GKGGAKRHRK VLRDNIQGIT KPAIRRLARR GGVKRISGLI YEETRGVLKV FLENVIRDAV TYTEHAKRKT VTAMDVVYAL KRQGRTLYGF GG

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Macromolecule #3: human histone H2A

MacromoleculeName: human histone H2A / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
SGRGKQGGKA RAKAKTRSSR AGLQFPVGRV HRLLRKGNYA ERVGAGAPVY LAAVLEYLTA EILELAGNAA RDNKKTRIIP RHLQLAIRND EELNKLLGKV TIAQGGVLPN IQAVLLPKKT ESHHKAKGK

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Macromolecule #4: human histone H2B

MacromoleculeName: human histone H2B / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
PEPAKSAPAP KKGSKKAVTK AQKKDGKKRK RSRKESYSVY VYKVLKQVHP DTGISSKAMG IMNSFVNDIF ERIAGEASRL AHYNKRSTIT SREIQTAVRL LLPGELAKHA VSEGTKAVTK YTSSK

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Macromolecule #7: Lysine-specific demethylase 2B (KDM2B)

MacromoleculeName: Lysine-specific demethylase 2B (KDM2B) / type: protein_or_peptide / ID: 7 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: GSEPEEERIR YSQRLRGTMR RRYEDDGISD DEIEGKRTFD LEEKLHTNKY NANFVTFMEG KDFNVEYIQR GGLRDPLIFK NSDGLGIKMP DPDFTVNDVK MCVGSRRMVD VMDVNTQKGI EMTMAQWTRY YETPEEEREK LYNVISLEFS HTRLENMVQR PSTVDFIDWV ...String:
GSEPEEERIR YSQRLRGTMR RRYEDDGISD DEIEGKRTFD LEEKLHTNKY NANFVTFMEG KDFNVEYIQR GGLRDPLIFK NSDGLGIKMP DPDFTVNDVK MCVGSRRMVD VMDVNTQKGI EMTMAQWTRY YETPEEEREK LYNVISLEFS HTRLENMVQR PSTVDFIDWV DNMWPRHLKE SQTESTNAIL EMQYPKVQKY CLMSVRGCYT DFHVDFGGTS VWYHIHQGGK VFWLIPPTAH NLELYENWLL SGKQGDIFLG DRVSDCQRIE LKQGYTFVIP SGWIHAVYTP TDTLVFGGNF LHSFNIPMQL KIYNIEDRTR VPNKFRYPFY YEMCWYVLER YVYCITNRSH LTKEFQKESL SMDLELNGLE SGNGDEEAVD REPRRLSSRR SVLTSPVANG VNLDYDGLGK TCRSLPSLKK TLAGDSSSDC SRGSHNGQVW DPQCAPRKDR QVHLTHFELE GLRCLVDKLE SLPLHKKCVP TGIEDEDALI ADVKILLEEL ANSDPKLALT GVPIVQWPKR DKLKFPTRPK VRVPTIPITK PHTMKPAPRL TPVRPAAASP IVSGARRRRV RCRKCKACVQ GECGVCHYCR DMKKFGGPGR MKQSCVLRQC LAPRLPHSVT CSLCGEVDQN EETQDFEKKL MECCICNEIV HPGCLQMDGE GLLNEELPNC WECPKCYQED SSEKAQHHHH HH

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Macromolecule #5: DNA 185-MER

MacromoleculeName: DNA 185-MER / type: dna / ID: 5 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
SequenceString:
ATCCCTATAC GCGGCCGCCC TGGAGAATCC CGGTGCCGAG GCCGCTCAAT TGGTCGTAGA CAGCTCTAGC ACCGCTTAAA CGCACGTACG CGCTGTCCCC CGCGTTTTAA CCGCCAAGGG GATTACTCCC TAGTCTCCAG GCACGTGTCA GATATATACA TCCTGTGCAT GTATTGAACA GCGAT

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Macromolecule #6: DNA 185-MER

MacromoleculeName: DNA 185-MER / type: dna / ID: 6 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
SequenceString:
ATCGCTGTTC AATACATGCA CAGGATGTAT ATATCTGACA CGTGCCTGGA GACTAGGGAG TAATCCCCTT GGCGGTTAAA ACGCGGGGGA CAGCGCGTAC GTGCGTTTAA GCGGTGCTAG AGCTGTCTAC GACCAATTGA GCGGCCTCGG CACCGGGATT CTCCAGGGCG GCCGCGTATA GGGAT

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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

Concentration1 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
10.0 mMC8H18N2O4SHEPES
30.0 mMNaClsodium chloride
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 15671 / Average electron dose: 45.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 45000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 3435247
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

7jo9

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 47729
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

7jo9

RefinementSpace: REAL

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