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- EMDB-22417: ORC-O1AAA: Human Origin Recognition Complex (ORC) with dynamic/un... -

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Basic information

Entry
Database: EMDB / ID: EMD-22417
TitleORC-O1AAA: Human Origin Recognition Complex (ORC) with dynamic/unresolved ORC2 WH
Map data
SampleORC-O1AAA
  • (Origin recognition complex subunit ...) x 5
  • (ligand) x 3
Function / homology
Function and homology information


origin recognition complex / inner kinetochore / nuclear origin of replication recognition complex / nuclear pre-replicative complex / mitotic DNA replication checkpoint signaling / DNA replication preinitiation complex / neural precursor cell proliferation / pre-replicative complex assembly involved in nuclear cell cycle DNA replication / DNA replication origin binding / regulation of DNA replication ...origin recognition complex / inner kinetochore / nuclear origin of replication recognition complex / nuclear pre-replicative complex / mitotic DNA replication checkpoint signaling / DNA replication preinitiation complex / neural precursor cell proliferation / pre-replicative complex assembly involved in nuclear cell cycle DNA replication / DNA replication origin binding / regulation of DNA replication / heterochromatin / regulation of transcription involved in G1/S transition of mitotic cell cycle / DNA replication initiation / pre-replicative complex assembly / chromosome, telomeric region / nuclear body / DNA replication / ATPase / centrosome / chromatin / nucleotide binding / chromatin binding / nucleolus / negative regulation of transcription by RNA polymerase II / DNA binding / membrane / nucleoplasm / ATP binding / metal ion binding / nucleus / cytosol
Bromo adjacent homology (BAH) domain / Origin recognition complex subunit 4, C-terminal / ATPase, AAA-type, core / Origin recognition complex, subunit 2 / Cdc6, C-terminal / Origin recognition complex subunit 4 / Origin recognition complex, subunit 1 / AAA+ ATPase domain / Origin recognition complex, subunit 5 / P-loop containing nucleoside triphosphate hydrolase ...Bromo adjacent homology (BAH) domain / Origin recognition complex subunit 4, C-terminal / ATPase, AAA-type, core / Origin recognition complex, subunit 2 / Cdc6, C-terminal / Origin recognition complex subunit 4 / Origin recognition complex, subunit 1 / AAA+ ATPase domain / Origin recognition complex, subunit 5 / P-loop containing nucleoside triphosphate hydrolase / Origin recognition complex, subunit 3 / ORC3, winged helix C-terminal / AAA lid domain / Orc1-like, AAA ATPase domain / Bromo adjacent homology (BAH) domain superfamily
Origin recognition complex subunit 5 / Origin recognition complex subunit 4 / Origin recognition complex subunit 1 / Origin recognition complex subunit 2 / Origin recognition complex subunit 3
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsJaremko MJ / Joshua-Tor L
Funding support United States, 3 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM45436 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)F32GM129923 United States
CitationJournal: Elife / Year: 2020
Title: The dynamic nature of the human origin recognition complex revealed through five cryoEM structures.
Authors: Matt J Jaremko / Kin Fan On / Dennis R Thomas / Bruce Stillman / Leemor Joshua-Tor /
Abstract: Genome replication is initiated from specific origin sites established by dynamic events. The Origin Recognition Complex (ORC) is necessary for orchestrating the initiation process by binding to ...Genome replication is initiated from specific origin sites established by dynamic events. The Origin Recognition Complex (ORC) is necessary for orchestrating the initiation process by binding to origin DNA, recruiting CDC6, and assembling the MCM replicative helicase on DNA. Here we report five cryoEM structures of the human ORC (HsORC) that illustrate the native flexibility of the complex. The absence of ORC1 revealed a compact, stable complex of ORC2-5. Introduction of ORC1 opens the complex into several dynamic conformations. Two structures revealed dynamic movements of the ORC1 AAA+ and ORC2 winged-helix domains that likely impact DNA incorporation into the ORC core. Additional twist and pinch motions were observed in an open ORC conformation revealing a hinge at the ORC5·ORC3 interface that may facilitate ORC binding to DNA. Finally, a structure of ORC was determined with endogenous DNA bound in the core revealing important differences between human and yeast origin recognition.
Validation ReportSummary, Full report, XML, About validation report
History
DepositionAug 9, 2020-
Header (metadata) releaseSep 2, 2020-
Map releaseSep 2, 2020-
UpdateSep 2, 2020-
Current statusSep 2, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.027
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.027
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7jpo
  • Surface level: 0.027
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_22417.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 320 pix.
= 342.4 Å
1.07 Å/pix.
x 320 pix.
= 342.4 Å
1.07 Å/pix.
x 320 pix.
= 342.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.07 Å
Density
Contour LevelBy AUTHOR: 0.027 / Movie #1: 0.027
Minimum - Maximum-0.07263519 - 0.16221082
Average (Standard dev.)0.0010934744 (±0.0033720883)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 342.40002 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.071.071.07
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z342.400342.400342.400
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ384384384
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.0730.1620.001

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Supplemental data

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Additional map: global refinement

Fileemd_22417_additional_1.map
Annotationglobal refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: focused refinement

Fileemd_22417_additional_2.map
Annotationfocused refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire ORC-O1AAA

EntireName: ORC-O1AAA
Details: 5 subunit complex with the ORC2 winged-helix domain in a dynamic/unresolved state
Number of components: 9

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Component #1: protein, ORC-O1AAA

ProteinName: ORC-O1AAA
Details: 5 subunit complex with the ORC2 winged-helix domain in a dynamic/unresolved state
Recombinant expression: No
MassTheoretical: 296.53 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #2: protein, Origin recognition complex subunit 1

ProteinName: Origin recognition complex subunit 1 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 44.310887 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #3: protein, Origin recognition complex subunit 2

ProteinName: Origin recognition complex subunit 2 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 66.063375 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #4: protein, Origin recognition complex subunit 3

ProteinName: Origin recognition complex subunit 3 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 82.436133 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #5: protein, Origin recognition complex subunit 4

ProteinName: Origin recognition complex subunit 4 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 50.443266 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #6: protein, Origin recognition complex subunit 5

ProteinName: Origin recognition complex subunit 5 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 50.298867 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #7: ligand, ADENOSINE-5'-TRIPHOSPHATE

LigandName: ADENOSINE-5'-TRIPHOSPHATE / Number of Copies: 3 / Recombinant expression: No
MassTheoretical: 0.507181 kDa

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Component #8: ligand, MAGNESIUM ION

LigandName: MAGNESIUM ION / Number of Copies: 3 / Recombinant expression: No
MassTheoretical: 2.430505 MDa

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Component #9: ligand, POTASSIUM ION

LigandName: POTASSIUM IONPotassium / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 3.909805 MDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.8 mg/mL / pH: 7.5
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 66 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 2.7 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 9068

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 160500
3D reconstructionSoftware: RELION / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement protocol: rigid body / Target criteria: Correlation Coefficient / Refinement space: REAL
Input PDB model: 5UJM, 5UJM, 5UJM, 5UJM, 5UJM
Chain ID: A, B, C, D, E
Modeling #2Refinement space: REAL
Input PDB model: 5UJM, 5UJM, 5UJM, 5UJM, 5UJM
Chain ID: A, B, C, D, E
Output model

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