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- EMDB-41839: Cryo-EM structure of yeast SWR1C subunit Swc5 bound to the nucleo... -

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Basic information

Entry
Database: EMDB / ID: EMD-41839
TitleCryo-EM structure of yeast SWR1C subunit Swc5 bound to the nucleosome, 3D class 0
Map dataMap of 3D class 0 (Fig. 6B, S1E)
Sample
  • Complex: Swc5 in complex with nucleosome core particle
    • Protein or peptide: Histone H3.2
    • Protein or peptide: Histone H4
    • Protein or peptide: Histone H2A.1
    • Protein or peptide: Histone H2B.1
    • Protein or peptide: SWR1-complex protein 5
    • DNA: Widom 601 forward DNA
    • DNA: Widom 601 reverse DNA
KeywordsChromatin / Remodeler / Complex / Histone / GENE REGULATION
Function / homology
Function and homology information


HATs acetylate histones / SIRT1 negatively regulates rRNA expression / Condensation of Prophase Chromosomes / Activated PKN1 stimulates transcription of AR (androgen receptor) regulated genes KLK2 and KLK3 / Assembly of the ORC complex at the origin of replication / Swr1 complex / HDACs deacetylate histones / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / Oxidative Stress Induced Senescence / replication fork protection complex ...HATs acetylate histones / SIRT1 negatively regulates rRNA expression / Condensation of Prophase Chromosomes / Activated PKN1 stimulates transcription of AR (androgen receptor) regulated genes KLK2 and KLK3 / Assembly of the ORC complex at the origin of replication / Swr1 complex / HDACs deacetylate histones / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / Oxidative Stress Induced Senescence / replication fork protection complex / RMTs methylate histone arginines / postreplication repair / RNA Polymerase I Promoter Escape / Estrogen-dependent gene expression / heterochromatin organization / Ub-specific processing proteases / nucleosomal DNA binding / structural constituent of chromatin / nucleosome / nucleosome assembly / chromatin organization / chromatin remodeling / protein heterodimerization activity / DNA repair / chromatin / regulation of DNA-templated transcription / negative regulation of transcription by RNA polymerase II / DNA binding / nucleoplasm / nucleus / cytosol
Similarity search - Function
BCNT-C domain / SWR1-complex protein 5/Craniofacial development protein 1/2 / Bucentaur or craniofacial development / Bucentaur C-terminal (BCNT-C) domain profile. / Histone H2B signature. / Histone H2B / Histone H2B / Histone H2A conserved site / Histone H2A signature. / Histone H2A, C-terminal domain ...BCNT-C domain / SWR1-complex protein 5/Craniofacial development protein 1/2 / Bucentaur or craniofacial development / Bucentaur C-terminal (BCNT-C) domain profile. / Histone H2B signature. / Histone H2B / Histone H2B / Histone H2A conserved site / Histone H2A signature. / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone H2A / Histone 2A / Histone H4, conserved site / Histone H4 signature. / Histone H4 / Histone H4 / CENP-T/Histone H4, histone fold / Centromere kinetochore component CENP-T histone fold / TATA box binding protein associated factor / TATA box binding protein associated factor (TAF), histone-like fold domain / Histone H3 signature 1. / Histone H3 signature 2. / Histone H3 / Histone H3/CENP-A / Histone H2A/H2B/H3 / Core histone H2A/H2B/H3/H4 / Histone-fold
Similarity search - Domain/homology
Histone H2B.1 / Histone H2A.1 / SWR1-complex protein 5 / Histone H4 / Histone H3.2
Similarity search - Component
Biological speciesXenopus laevis (African clawed frog) / Saccharomyces cerevisiae (brewer's yeast) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsEek P / Tan S
Funding support United States, Estonia, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM122519 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM127034 United States
Estonian Research CouncilPUTJD906 Estonia
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)U24GM129547 United States
Citation
Journal: Elife / Year: 2024
Title: Dual engagement of the nucleosomal acidic patches is essential for deposition of histone H2A.Z by SWR1C.
Authors: Alexander S Baier / Nathan Gioacchini / Priit Eek / Erik M Leith / Song Tan / Craig L Peterson /
Abstract: The yeast SWR1C chromatin remodeling enzyme catalyzes the ATP-dependent exchange of nucleosomal histone H2A for the histone variant H2A.Z, a key variant involved in a multitude of nuclear functions. ...The yeast SWR1C chromatin remodeling enzyme catalyzes the ATP-dependent exchange of nucleosomal histone H2A for the histone variant H2A.Z, a key variant involved in a multitude of nuclear functions. How the 14-subunit SWR1C engages the nucleosomal substrate remains largely unknown. Studies on the ISWI, CHD1, and SWI/SNF families of chromatin remodeling enzymes have demonstrated key roles for the nucleosomal acidic patch for remodeling activity, however a role for this nucleosomal epitope in nucleosome editing by SWR1C has not been tested. Here, we employ a variety of biochemical assays to demonstrate an essential role for the acidic patch in the H2A.Z exchange reaction. Utilizing asymmetrically assembled nucleosomes, we demonstrate that the acidic patches on each face of the nucleosome are required for SWR1C-mediated dimer exchange, suggesting SWR1C engages the nucleosome in a 'pincer-like' conformation, engaging both patches simultaneously. Loss of a single acidic patch results in loss of high affinity nucleosome binding and nucleosomal stimulation of ATPase activity. We identify a conserved arginine-rich motif within the Swc5 subunit that binds the acidic patch and is key for dimer exchange activity. In addition, our cryoEM structure of a Swc5-nucleosome complex suggests that promoter proximal, histone H2B ubiquitylation may regulate H2A.Z deposition. Together these findings provide new insights into how SWR1C engages its nucleosomal substrate to promote efficient H2A.Z deposition.
#1: Journal: Elife / Year: 2024
Title: Dual engagement of the nucleosomal acidic patches is essential for deposition of histone H2A.Z by SWR1C
Authors: Baier AS / Gioacchini N / Eek P / Leith EM / Song T / Peterson CL
History
DepositionSep 5, 2023-
Header (metadata) releaseMay 29, 2024-
Map releaseMay 29, 2024-
UpdateJun 12, 2024-
Current statusJun 12, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41839.png

Downloads & links

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Map

FileDownload / File: emd_41839.map.gz / Format: CCP4 / Size: 38.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMap of 3D class 0 (Fig. 6B, S1E)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.18 Å/pix.
x 216 pix.
= 255.312 Å		1.18 Å/pix.
x 216 pix.
= 255.312 Å		1.18 Å/pix.
x 216 pix.
= 255.312 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.182 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.06
Minimum - Maximum-0.31823304 - 0.6708708
Average (Standard dev.)0.000025005054 (±0.02338448)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions216216216
Spacing216216216
CellA=B=C: 255.31201 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_41839_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B of class 0

Fileemd_41839_half_map_1.map
AnnotationHalf map B of class 0
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A of class 0

Fileemd_41839_half_map_2.map
AnnotationHalf map A of class 0
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Swc5 in complex with nucleosome core particle

EntireName: Swc5 in complex with nucleosome core particle
Components
  • Complex: Swc5 in complex with nucleosome core particle
    • Protein or peptide: Histone H3.2
    • Protein or peptide: Histone H4
    • Protein or peptide: Histone H2A.1
    • Protein or peptide: Histone H2B.1
    • Protein or peptide: SWR1-complex protein 5
    • DNA: Widom 601 forward DNA
    • DNA: Widom 601 reverse DNA

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Supramolecule #1: Swc5 in complex with nucleosome core particle

SupramoleculeName: Swc5 in complex with nucleosome core particle / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Xenopus laevis (African clawed frog)

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Macromolecule #1: Histone H3.2

MacromoleculeName: Histone H3.2 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
ARTKQTARKS TGGKAPRKQL ATKAARKSAP ATGGVKKPHR YRPGTVALRE IRRYQKSTEL LIRKLPFQRL VREIAQDFKT DLRFQSSAVM ALQEASEAYL VALFEDTNLC AIHAKRVTIM PKDIQLARRI RGERA

UniProtKB: Histone H3.2

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Macromolecule #2: Histone H4

MacromoleculeName: Histone H4 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MSGRGKGGKG LGKGGAKRHR KVLRDNIQGI TKPAIRRLAR RGGVKRISGL IYEETRGVLK VFLENVIRDA VTYTEHAKRK TVTAMDVVYA LKRQGRTLYG FGG

UniProtKB: Histone H4

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Macromolecule #3: Histone H2A.1

MacromoleculeName: Histone H2A.1 / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
SGGKGGKAGS AAKASQSRSA KAGLTFPVGR VHRLLRRGNY AQRIGSGAPV YLTAVLEYLA AEILELAGNA ARDNKKTRII PRHLQLAIRN DDELNKLLGN VTIAQGGVLP NIHQNLLPKK SAKATKASQE L

UniProtKB: Histone H2A.1

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Macromolecule #4: Histone H2B.1

MacromoleculeName: Histone H2B.1 / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
SAKAEKKPAS KAPAEKKPAA KKTSTSTDGK KRSKARKETY SSYIYKVLKQ THPDTGISQK SMSILNSFVN DIFERIATEA SKLAAYNKKS TISAREIQTA VRLILPGELA KHAVSEGTRA VTKYSSSTQA

UniProtKB: Histone H2B.1

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Macromolecule #5: SWR1-complex protein 5

MacromoleculeName: SWR1-complex protein 5 / type: protein_or_peptide / ID: 5 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: RGSHHHHHHG SKTRRARQAE EEYAKTHKYE SLTVESIPAK VNSIWEELQE ASKNRLLSSS GKVGSVLDGS KEARSTTAAQ QEDKILIERN YKFAGETVHE KKWVSRSSAE GQEYLNSLKF KQQAPAAPVQ LEKAVRTKSN ESRQHLRRPL KRPPLLEQII SGGLRPKLTT ...String:
RGSHHHHHHG SKTRRARQAE EEYAKTHKYE SLTVESIPAK VNSIWEELQE ASKNRLLSSS GKVGSVLDGS KEARSTTAAQ QEDKILIERN YKFAGETVHE KKWVSRSSAE GQEYLNSLKF KQQAPAAPVQ LEKAVRTKSN ESRQHLRRPL KRPPLLEQII SGGLRPKLTT LEKSQLDWAS YVDRAGLNDE LVLHNKDGFL ARQEFLQRVG SAEDERYKEL RRQQLAQQLQ QDNEAS

UniProtKB: SWR1-complex protein 5

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Macromolecule #6: Widom 601 forward DNA

MacromoleculeName: Widom 601 forward DNA / type: dna / ID: 6 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
SequenceString:
ATCGAGAATC CCGGTGCCGA GGCCGCTCAA TTGGTCGTAG ACAGCTCTAG CACCGCTTAA ACGCACGTAC GCGCTGTCCC CCGCGTTTTA ACCGCCAAGG GGATTACTCC CTAGTCTCCA GGCACGTGTC AGATATATAC ATCCGAT

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Macromolecule #7: Widom 601 reverse DNA

MacromoleculeName: Widom 601 reverse DNA / type: dna / ID: 7 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
SequenceString:
ATCGGATGTA TATATCTGAC ACGTGCCTGG AGACTAGGGA GTAATCCCCT TGGCGGTTAA AACGCGGGGG ACAGCGCGTA CGTGCGTTTA AGCGGTGCTA GAGCTGTCTA CGACCAATTG AGCGGCCTCG GCACCGGGAT TCTCGAT

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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

Concentration0.9 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
10.0 mMHEPES2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid
50.0 mMNaClSodium chloridesodium chloride
1.0 mMDTTdithiothreitol
0.1 mMPMSFphenylmethylsulfonyl fluoridePMSF
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: AIR / Details: 15 mA with PELCO easiGlow
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
DetailsChemically cross-linked with glutaraldehyde using the GraFix method.

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.8000000000000003 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 29000
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 13855 / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 4564529
Startup modelType of model: INSILICO MODEL / In silico model: Ab-initio model
Details: Created with CryoSPARC Ab-Initio Reconstruction into 3 classes.
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v4.1.2) / Software - details: Ab-Initio Reconstruction
Final 3D classificationNumber classes: 10 / Avg.num./class: 59000 / Software - Name: cryoSPARC (ver. v4.2.0) / Software - details: 3D Classification
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v4.2.1) / Software - details: Non-uniform Refinement
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. v4.2.1) / Software - details: Non-uniform refinement / Number images used: 63105
FSC plot (resolution estimation)

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