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- EMDB-56897: sCMGE assembled on ARS1 DNA with RPA and no Sld2 -

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Basic information

Entry
Database: EMDB / ID: EMD-56897
TitlesCMGE assembled on ARS1 DNA with RPA and no Sld2
Map data3D Refinement of sCMGE on dsDNA assembled in the absence of Sld2 Relion Unsharpened map
Sample
  • Complex: sCMGE assembled on ARS1 DNA with RPA and no Sld2
KeywordsMacromolecular Complex DNA ATPase Helicase MCM2-7 / REPLICATION
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsPuehringer T / Palm G / Costa A
Funding support United Kingdom, European Union, Germany, 6 items
OrganizationGrant numberCountry
The Francis Crick Institute United Kingdom
Wellcome Trust United Kingdom
Medical Research Council (MRC, United Kingdom) United Kingdom
Cancer Research UK United Kingdom
European Research Council (ERC)European Union
Boehringer Ingelheim Fonds (BIF) Germany
CitationJournal: Nature / Year: 2026
Title: Structure of the pre-initiation complex explains CMGE biogenesis.
Authors: Thomas Pühringer / Berta Canal / Giacomo Palm / Agata Butryn / Emma C Couves / Oliver Willhoft / Jacob S Lewis / John F X Diffley / Alessandro Costa /
Abstract: When cells enter S phase, bidirectional DNA replication is initiated through the kinase-regulated recruitment of three activators (Cdc45, GINS and Pol ε) to a duplex-DNA-loaded double hexamer of ...When cells enter S phase, bidirectional DNA replication is initiated through the kinase-regulated recruitment of three activators (Cdc45, GINS and Pol ε) to a duplex-DNA-loaded double hexamer of minichromosome maintenance (MCM) ATPases. Together, these proteins form two CMGE helicases that establish divergent replication forks as they become separated. Here, to gain an understanding of CMGE biogenesis, we reconstituted the pre-initiation complex with purified yeast proteins. The cryo-electron-microscopy structure shows a set of firing factors caught in the act of assembling two symmetrical CMGEs. We show how stepwise complex formation reshapes MCM in preparation for DNA opening, and we explain how ATP promotes firing-factor ejection and CMGE maturation. We find that although Sld2 facilitates the recruitment of GINS to MCM, as expected, it also aids the efficient separation of the CMGE dimer, and is essential for the ejection of the lagging strand from MCM. These findings have direct implications for our understanding of the metazoan Sld2 orthologue, RECQL4, and point to a replication-fork establishment mechanism that is conserved across eukaryotes.
History
DepositionFeb 23, 2026-
Header (metadata) releaseMay 13, 2026-
Map releaseMay 13, 2026-
UpdateJul 1, 2026-
Current statusJul 1, 2026Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_56897.png

Downloads & links

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Map

FileDownload / File: emd_56897.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation3D Refinement of sCMGE on dsDNA assembled in the absence of Sld2 Relion Unsharpened map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.95 Å/pix.
x 512 pix.
= 486.4 Å		0.95 Å/pix.
x 512 pix.
= 486.4 Å		0.95 Å/pix.
x 512 pix.
= 486.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.95 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0045
Minimum - Maximum-0.010657752 - 0.02813799
Average (Standard dev.)0.000045268156 (±0.00081819145)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 486.4 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: 3D Refinement of sCMGE on dsDNA assembled in...

Fileemd_56897_additional_1.map
Annotation3D Refinement of sCMGE on dsDNA assembled in the absence of Sld2 Relion Density-modified map (EMReady)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: 3D Refinement of sCMGE on dsDNA assembled in...

Fileemd_56897_half_map_1.map
Annotation3D Refinement of sCMGE on dsDNA assembled in the absence of Sld2 Relion Half map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: 3D Refinement of sCMGE on dsDNA assembled in...

Fileemd_56897_half_map_2.map
Annotation3D Refinement of sCMGE on dsDNA assembled in the absence of Sld2 Relion Half map 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 : sCMGE assembled on ARS1 DNA with RPA and no Sld2

EntireName: sCMGE assembled on ARS1 DNA with RPA and no Sld2
Components
  • Complex: sCMGE assembled on ARS1 DNA with RPA and no Sld2

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Supramolecule #1: sCMGE assembled on ARS1 DNA with RPA and no Sld2

SupramoleculeName: sCMGE assembled on ARS1 DNA with RPA and no Sld2 / type: complex / ID: 1 / Parent: 0
Macromolecule list: #6, #13-#14, #16, #7-#10, #12, #2, #17, #4, #3, #1, #11, #15, #5
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)

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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

BufferpH: 7.5
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Number grids imaged: 1 / Number real images: 70337 / Average electron dose: 42.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.4000000000000001 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2390783
CTF correctionType: NONE
Startup modelType of model: INSILICO MODEL / In silico model: cryoSPARC ab initio volume
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 5) / Number images used: 72370
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 5)
FSC plot (resolution estimation)

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