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- EMDB-0091: Structure of a yeast closed complex with distorted DNA (core CCdist) -

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Entry
Database: EMDB / ID: 0091
TitleStructure of a yeast closed complex with distorted DNA (core CCdist)
Map dataLocally filtered map
SamplePre-initiation complexPreinitiation complex
  • Polymerase
  • TIFs
  • nucleic acidsNucleic acid
  • (DNA-directed RNA polymerase II subunit ...Polymerase) x 7
  • (DNA-directed RNA polymerases I, II, and III subunit ...RNA polymerase) x 5
  • Transcription initiation factor IIB
  • (nucleic-acidNucleic acid) x 2
  • TATA-box-binding protein
  • (Transcription initiation factor IIF subunit ...) x 2
  • (Transcription initiation factor IIA ...) x 2
  • (Transcription initiation factor IIE subunit ...) x 2
  • (ligand) x 2
Function / homologyRNA polymerase subunit, RPB6/omega / DNA-directed RNA polymerase M, 15kDa subunit, conserved site / Zinc finger, TFIIB-type / Transcription factor TFIIB, cyclin-like domain / Cyclin-like / DNA-directed RNA polymerase RPB5 subunit, eukaryote/virus / RNA polymerase Rpb2, OB-fold / DNA-directed RNA polymerase, subunit 2 / Transcription initiation factor IIA, gamma subunit, C-terminal / Transcription initiation factor IIA, gamma subunit, N-terminal ...RNA polymerase subunit, RPB6/omega / DNA-directed RNA polymerase M, 15kDa subunit, conserved site / Zinc finger, TFIIB-type / Transcription factor TFIIB, cyclin-like domain / Cyclin-like / DNA-directed RNA polymerase RPB5 subunit, eukaryote/virus / RNA polymerase Rpb2, OB-fold / DNA-directed RNA polymerase, subunit 2 / Transcription initiation factor IIA, gamma subunit, C-terminal / Transcription initiation factor IIA, gamma subunit, N-terminal / Transcription initiation factor TFIIE, beta subunit / Transcription factor TFE/TFIIEalpha HTH domain / RNA polymerase, subunit H/Rpb5, conserved site / Nucleic acid-binding, OB-fold / Archaeal RpoH /eukaryotic RPB5 RNA polymerase subunit / DNA-directed RNA polymerase, 14-18kDa subunit, conserved site / RNA polymerases, subunit N, zinc binding site / RNA-binding domain, S1 / Transcription factor TFIIB, conserved site / RNA polymerase subunit RPB10 / TFIIEalpha/SarR/Rpc3 HTH domain / DNA-directed RNA polymerase, subunit RPB6 / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / TATA-box binding protein, conserved site / TATA-box binding protein, eukaryotic / Zinc finger, RING/FYVE/PHD-type / TBP domain superfamily / RPB5-like RNA polymerase subunit superfamily / RNA polymerase, beta subunit, protrusion / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 7 / RNA polymerase Rpb1, domain 6 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain / RNA polymerase, beta subunit, conserved site / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 2 / RNA polymerase Rpb2, domain 3 / DNA-directed RNA polymerase subunit/transcription factor S / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 5 / DNA-directed RNA polymerase Rpb11, 13-16kDa subunit, conserved site / Transcription initiation factor IIF, alpha subunit / DNA-directed RNA polymerase, RBP11-like dimerisation domain / Transcription factor IIA, helical / Transcription factor IIA, beta-barrel / HRDC-like superfamily / Transcription Factor IIF, Rap30/Rap74, interaction / DNA-directed RNA polymerase, insert domain / DNA-directed RNA polymerase, RpoA/D/Rpb3-type / Pol II subunit B9, C-terminal zinc ribbon / RPB6/omega subunit-like superfamily / RNA polymerase archaeal subunit P/eukaryotic subunit RPABC4 / SHS2 domain found in N terminus of Rpb7p/Rpc25p/MJ0397 / RNA polymerases M/15 Kd subunit / TFIIE beta subunit core domain / Transcription initiation factor IIA, gamma subunit, helical domain / TFIIF, beta subunit HTH domain / Transcription initiation factor IIA, gamma subunit / Transcription factor IIA, alpha/beta subunit / DNA directed RNA polymerase, 7 kDa subunit / RNA polymerase Rpb8 / RNA polymerase Rpb5, N-terminal domain / RNA polymerase Rpb4 / RNA polymerase Rpb2, domain 7 / RNA polymerases N / 8 kDa subunit / RNA polymerase Rpb2, domain 2 / RNA polymerase beta subunit / RNA polymerase Rpb2, domain 3 / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 5 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 7 / RNA polymerase Rpb1, domain 6 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / TFIIE alpha subunit / RNA polymerase Rpb3/Rpb11 dimerisation domain / Winged helix-like DNA-binding domain superfamily / Rpb4/RPC9 superfamily / Winged helix DNA-binding domain superfamily / RNA polymerase, RBP11-like subunit / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase Rpb5, N-terminal domain superfamily / RNA polymerase Rpb7-like, N-terminal domain superfamily / Cyclin-like superfamily / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase RBP11 / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase Rpb1, domain 7 superfamily / RNA polymerase Rpb6 / DNA-directed RNA polymerase subunit Rpb5-like / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / Transcription factor E / Transcription factor TFIID (or TATA-binding protein, TBP) / Transcription factor TFIIB repeat
Function and homology information
SourceSaccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) / Baker's yeast (baker's yeast)
Methodsingle particle reconstruction / cryo EM / 4.8 Å resolution
AuthorsDienemann C / Schwalb B / Schilbach S / Cramer P
CitationJournal: Mol. Cell / Year: 2018
Title: Promoter Distortion and Opening in the RNA Polymerase II Cleft.
Authors: Christian Dienemann / Björn Schwalb / Sandra Schilbach / Patrick Cramer
Abstract: Transcription initiation requires opening of promoter DNA in the RNA polymerase II (Pol II) pre-initiation complex (PIC), but it remains unclear how this is achieved. Here we report the cryo-electron ...Transcription initiation requires opening of promoter DNA in the RNA polymerase II (Pol II) pre-initiation complex (PIC), but it remains unclear how this is achieved. Here we report the cryo-electron microscopic (cryo-EM) structure of a yeast PIC that contains underwound, distorted promoter DNA in the closed Pol II cleft. The DNA duplex axis is offset at the upstream edge of the initially melted DNA region (IMR) where DNA opening begins. Unstable IMRs are found in a subset of yeast promoters that we show can still initiate transcription after depletion of the transcription factor (TF) IIH (TFIIH) translocase Ssl2 (XPB in human) from the nucleus in vivo. PIC-induced DNA distortions may thus prime the IMR for melting and may explain how unstable IMRs that are predicted in promoters of Pol I and Pol III can open spontaneously. These results suggest that DNA distortion in the polymerase cleft is a general mechanism that contributes to promoter opening.
Validation ReportPDB-ID: 6gyl

SummaryFull reportAbout validation report
DateDeposition: Jun 30, 2018 / Header (metadata) release: Aug 22, 2018 / Map release: Dec 5, 2018 / Last update: Dec 5, 2018

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0028
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.0028
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6gyl
  • Surface level: 0.0028
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_0091.map.gz (map file in CCP4 format, 108001 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
300 pix
1.05 Å/pix.
= 315. Å
300 pix
1.05 Å/pix.
= 315. Å
300 pix
1.05 Å/pix.
= 315. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.05 Å
Density
Contour Level:0.0028 (by author), 0.0028 (movie #1):
Minimum - Maximum-0.010854901 - 0.021589203
Average (Standard dev.)-0.000029907023 (0.0007890197)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions300300300
Origin0.00.00.0
Limit299.0299.0299.0
Spacing300300300
CellA=B=C: 315.0 Å
α=β=γ: 90.0 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.051.051.05
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z315.000315.000315.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ450450450
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-0.0110.022-0.000

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Supplemental data

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Sample components

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Entire Pre-initiation complex

EntireName: Pre-initiation complex / Number of components: 28

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Component #1: protein, Pre-initiation complex

ProteinName: Pre-initiation complexPreinitiation complex / Recombinant expression: No

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Component #2: protein, Polymerase

ProteinName: Polymerase / Recombinant expression: No
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)

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Component #3: protein, TIFs

ProteinName: TIFs / Recombinant expression: No
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #4: protein, nucleic acids

ProteinName: nucleic acidsNucleic acid / Recombinant expression: No
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: synthetic construct (others)

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Component #5: protein, DNA-directed RNA polymerase II subunit RPB1

ProteinName: DNA-directed RNA polymerase II subunit RPB1Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 191.821578 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #6: protein, DNA-directed RNA polymerase II subunit RPB2

ProteinName: DNA-directed RNA polymerase II subunit RPB2Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 138.937297 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #7: protein, DNA-directed RNA polymerase II subunit RPB3

ProteinName: DNA-directed RNA polymerase II subunit RPB3Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 35.330457 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #8: protein, DNA-directed RNA polymerase II subunit RPB4

ProteinName: DNA-directed RNA polymerase II subunit RPB4Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 25.451191 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #9: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC1

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC1RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 25.117094 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #10: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC2

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC2RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 17.931834 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #11: protein, DNA-directed RNA polymerase II subunit RPB7

ProteinName: DNA-directed RNA polymerase II subunit RPB7Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 19.081053 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #12: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC3

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC3RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 16.525363 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #13: protein, DNA-directed RNA polymerase II subunit RPB9

ProteinName: DNA-directed RNA polymerase II subunit RPB9Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 14.308161 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #14: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC5

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC5RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 8.290732 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #15: protein, DNA-directed RNA polymerase II subunit RPB11

ProteinName: DNA-directed RNA polymerase II subunit RPB11Polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 13.633493 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #16: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC4

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC4RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 7.729969 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #17: protein, Transcription initiation factor IIB

ProteinName: Transcription initiation factor IIB / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 38.25734 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #18: nucleic-acid, GAT1 promoter DNA

Nucleic-acidName: GAT1 promoter DNA / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DT)(DG)(DC)(DC)(DC)(DG)(DG)(DC)(DC)(DC) (DA)(DG)(DC)(DC)(DA)(DC)(DA)(DT)(DA)(DT) (DA)(DT)(DA)(DT)(DA)(DG)(DG)(DT)(DG)(DT) (DG)(DT)(DG)(DC)(DC)(DA)(DC)(DT)(DC)(DC) (DC)(DG)(DG)(DC)(DC)(DC)(DC)(DG)(DG)(DT) (DA)(DT)(DT)(DA)(DG)(DC)
MassTheoretical: 17.129941 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)

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Component #19: protein, TATA-box-binding protein

ProteinName: TATA-box-binding protein / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 27.042275 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #20: protein, Transcription initiation factor IIF subunit alpha

ProteinName: Transcription initiation factor IIF subunit alpha / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 82.32057 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #21: protein, Transcription initiation factor IIF subunit beta

ProteinName: Transcription initiation factor IIF subunit beta / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 46.684492 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #22: nucleic-acid, GAT1 promoter DNA

Nucleic-acidName: GAT1 promoter DNA / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DG)(DC)(DT)(DA)(DA)(DT)(DA)(DC)(DC)(DG) (DG)(DG)(DG)(DC)(DC)(DG)(DG)(DG)(DA)(DG) (DT)(DG)(DG)(DC)(DA)(DC)(DA)(DC)(DA)(DC) (DC)(DT)(DA)(DT)(DA)(DT)(DA)(DT)(DA)(DT) (DG)(DT)(DG)(DG)(DC)(DT)(DG)(DG)(DG)(DC) (DC)(DG)(DG)(DG)(DC)(DA)
MassTheoretical: 17.388111 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)

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Component #23: protein, Transcription initiation factor IIA large subunit,Transc...

ProteinName: Transcription initiation factor IIA large subunit,Transcription initiation factor IIA large subunit
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 19.432029 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #24: protein, Transcription initiation factor IIA subunit 2

ProteinName: Transcription initiation factor IIA subunit 2 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 14.431131 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #25: protein, Transcription initiation factor IIE subunit alpha,Tfa1,Tfa1

ProteinName: Transcription initiation factor IIE subunit alpha,Tfa1,Tfa1
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 57.538461 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #26: protein, Transcription initiation factor IIE subunit beta

ProteinName: Transcription initiation factor IIE subunit beta / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 37.050434 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #27: ligand, ZINC ION

LigandName: ZINC ION / Number of Copies: 10 / Recombinant expression: No
MassTheoretical: 6.540905 MDa

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Component #28: ligand, MAGNESIUM ION

LigandName: MAGNESIUM ION / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 2.430505 MDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionpH: 7
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 37 e/Å2 / Illumination mode: OTHER
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 QUANTUM (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 38000
3D reconstructionResolution: 4.8 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Output model

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