+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-13494 | ||||||||||||
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Title | H. sapiens replisome-CUL2/LRR1 complex | ||||||||||||
Map data | Locally filtered consensus refinement | ||||||||||||
Sample |
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Keywords | Genome stability / DNA replication / Ubiquitination / termination / replisome / cryo-EM / CMG / Cul2LRR1 / REPLICATION / DNA / polymerase / helicase | ||||||||||||
Function / homology | Function and homology information cellular response to bleomycin / DNA secondary structure binding / detection of abiotic stimulus / replication fork arrest / regulation of nuclear cell cycle DNA replication / Switching of origins to a post-replicative state / Unwinding of DNA / cell cycle phase transition / cellular response to cisplatin / DNA replication initiation ...cellular response to bleomycin / DNA secondary structure binding / detection of abiotic stimulus / replication fork arrest / regulation of nuclear cell cycle DNA replication / Switching of origins to a post-replicative state / Unwinding of DNA / cell cycle phase transition / cellular response to cisplatin / DNA replication initiation / epsilon DNA polymerase complex / DNA strand elongation involved in mitotic DNA replication / GINS complex / mitotic DNA replication preinitiation complex assembly / nuclear origin of replication recognition complex / cellular response to hydroxyurea / anaphase-promoting complex binding / cullin-RING-type E3 NEDD8 transferase / alpha DNA polymerase:primase complex / mitotic DNA replication / cellular response to chemical stress / NEDD8 transferase activity / cullin-RING ubiquitin ligase complex / CMG complex / DNA replication checkpoint signaling / Cul7-RING ubiquitin ligase complex / ubiquitin-dependent protein catabolic process via the C-end degron rule pathway / nucleotide-excision repair, DNA gap filling / single-stranded DNA 3'-5' DNA exonuclease activity / DNA replication preinitiation complex / target-directed miRNA degradation / MCM complex / regulation of phosphorylation / Loss of Function of FBXW7 in Cancer and NOTCH1 Signaling / replication fork protection complex / elongin complex / DNA replication proofreading / VCB complex / mitotic DNA replication checkpoint signaling / double-strand break repair via break-induced replication / positive regulation of protein autoubiquitination / protein neddylation / mitotic DNA replication initiation / regulation of DNA-templated DNA replication initiation / mitotic intra-S DNA damage checkpoint signaling / NEDD8 ligase activity / Cul5-RING ubiquitin ligase complex / positive regulation of double-strand break repair / negative regulation of response to oxidative stress / DNA strand elongation involved in DNA replication / ubiquitin-ubiquitin ligase activity / Hydrolases; Acting on ester bonds; Exodeoxyribonucleases producing 5'-phosphomonoesters / inner cell mass cell proliferation / SCF ubiquitin ligase complex / Cul2-RING ubiquitin ligase complex / Cul4A-RING E3 ubiquitin ligase complex / negative regulation of type I interferon production / ubiquitin ligase complex scaffold activity / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / Cul4B-RING E3 ubiquitin ligase complex / Cul3-RING ubiquitin ligase complex / activation of protein kinase activity / branching morphogenesis of an epithelial tube / DNA synthesis involved in DNA repair / cochlea development / G1/S-Specific Transcription / leading strand elongation / DNA unwinding involved in DNA replication / Apoptotic cleavage of cellular proteins / Prolactin receptor signaling / replication fork processing / nuclear replication fork / protein monoubiquitination / mitotic G2 DNA damage checkpoint signaling / DNA replication origin binding / Pausing and recovery of Tat-mediated HIV elongation / Tat-mediated HIV elongation arrest and recovery / cullin family protein binding / HIV elongation arrest and recovery / Pausing and recovery of HIV elongation / positive regulation of double-strand break repair via homologous recombination / PCNA-Dependent Long Patch Base Excision Repair / Activation of the pre-replicative complex / DNA replication initiation / Tat-mediated elongation of the HIV-1 transcript / error-prone translesion synthesis / embryonic organ development / Formation of HIV-1 elongation complex containing HIV-1 Tat / cellular response to interleukin-4 / Formation of HIV elongation complex in the absence of HIV Tat / protein K48-linked ubiquitination / Activation of ATR in response to replication stress / RNA Polymerase II Transcription Elongation / Nuclear events stimulated by ALK signaling in cancer / Formation of RNA Pol II elongation complex / response to UV / RNA Polymerase II Pre-transcription Events / base-excision repair, gap-filling / DNA helicase activity / positive regulation of TORC1 signaling Similarity search - Function | ||||||||||||
Biological species | Homo sapiens (human) | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.8 Å | ||||||||||||
Authors | Jones MJ / Yeeles JTP | ||||||||||||
Funding support | United Kingdom, 3 items
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Citation | Journal: Nature / Year: 2021 Title: A conserved mechanism for regulating replisome disassembly in eukaryotes. Authors: Michael Jenkyn-Bedford / Morgan L Jones / Yasemin Baris / Karim P M Labib / Giuseppe Cannone / Joseph T P Yeeles / Tom D Deegan / Abstract: Replisome disassembly is the final step of eukaryotic DNA replication and is triggered by ubiquitylation of the CDC45-MCM-GINS (CMG) replicative helicase. Despite being driven by evolutionarily ...Replisome disassembly is the final step of eukaryotic DNA replication and is triggered by ubiquitylation of the CDC45-MCM-GINS (CMG) replicative helicase. Despite being driven by evolutionarily diverse E3 ubiquitin ligases in different eukaryotes (SCF in budding yeast, CUL2 in metazoa), replisome disassembly is governed by a common regulatory principle, in which ubiquitylation of CMG is suppressed before replication termination, to prevent replication fork collapse. Recent evidence suggests that this suppression is mediated by replication fork DNA. However, it is unknown how SCF and CUL2 discriminate terminated from elongating replisomes, to selectively ubiquitylate CMG only after termination. Here we used cryo-electron microscopy to solve high-resolution structures of budding yeast and human replisome-E3 ligase assemblies. Our structures show that the leucine-rich repeat domains of Dia2 and LRR1 are structurally distinct, but bind to a common site on CMG, including the MCM3 and MCM5 zinc-finger domains. The LRR-MCM interaction is essential for replisome disassembly and, crucially, is occluded by the excluded DNA strand at replication forks, establishing the structural basis for the suppression of CMG ubiquitylation before termination. Our results elucidate a conserved mechanism for the regulation of replisome disassembly in eukaryotes, and reveal a previously unanticipated role for DNA in preserving replisome integrity. | ||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_13494.map.gz | 140.1 MB | EMDB map data format | |
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Header (meta data) | emd-13494-v30.xml emd-13494.xml | 60.9 KB 60.9 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_13494_fsc.xml | 18.3 KB | Display | FSC data file |
Images | emd_13494.png | 107.6 KB | ||
Masks | emd_13494_msk_1.map | 244.1 MB | Mask map | |
Filedesc metadata | emd-13494.cif.gz | 17.3 KB | ||
Others | emd_13494_additional_1.map.gz emd_13494_additional_2.map.gz emd_13494_half_map_1.map.gz emd_13494_half_map_2.map.gz | 123.3 MB 230.4 MB 226.9 MB 226.9 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-13494 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13494 | HTTPS FTP |
-Validation report
Summary document | emd_13494_validation.pdf.gz | 965.1 KB | Display | EMDB validaton report |
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Full document | emd_13494_full_validation.pdf.gz | 964.7 KB | Display | |
Data in XML | emd_13494_validation.xml.gz | 21.5 KB | Display | |
Data in CIF | emd_13494_validation.cif.gz | 28.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13494 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13494 | HTTPS FTP |
-Related structure data
Related structure data | 7ploMC 7pmkC 7pmnC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_13494.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Locally filtered consensus refinement | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.072 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_13494_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: consensus refinement
File | emd_13494_additional_1.map | ||||||||||||
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Annotation | consensus refinement | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: consensus refinement sharpened
File | emd_13494_additional_2.map | ||||||||||||
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Annotation | consensus refinement sharpened | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: consensus refinement half 1
File | emd_13494_half_map_1.map | ||||||||||||
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Annotation | consensus refinement half 1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: consensus refinement half 2
File | emd_13494_half_map_2.map | ||||||||||||
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Annotation | consensus refinement half 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : Human replisome engaged with CUL2-LRR1
+Supramolecule #1: Human replisome engaged with CUL2-LRR1
+Macromolecule #1: DNA replication licensing factor MCM2
+Macromolecule #2: DNA replication licensing factor MCM3
+Macromolecule #3: DNA replication licensing factor MCM4
+Macromolecule #4: DNA replication licensing factor MCM5
+Macromolecule #5: DNA replication licensing factor MCM6
+Macromolecule #6: DNA replication licensing factor MCM7
+Macromolecule #7: DNA polymerase epsilon subunit 2
+Macromolecule #8: DNA polymerase epsilon catalytic subunit A
+Macromolecule #9: Cell division control protein 45 homolog
+Macromolecule #10: DNA replication complex GINS protein PSF1
+Macromolecule #11: DNA replication complex GINS protein PSF2
+Macromolecule #12: DNA replication complex GINS protein PSF3
+Macromolecule #13: DNA replication complex GINS protein SLD5
+Macromolecule #14: WD repeat and HMG-box DNA-binding protein 1
+Macromolecule #15: Protein timeless homolog
+Macromolecule #16: TIMELESS-interacting protein
+Macromolecule #19: Leucine-rich repeat protein 1
+Macromolecule #20: Elongin-B
+Macromolecule #21: Claspin
+Macromolecule #22: Elongin-C
+Macromolecule #23: Cullin-2
+Macromolecule #24: E3 ubiquitin-protein ligase RBX1
+Macromolecule #17: Leading strand DNA
+Macromolecule #18: Lagging strand DNA
+Macromolecule #25: ZINC ION
+Macromolecule #26: MAGNESIUM ION
+Macromolecule #27: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER
+Macromolecule #28: SULFATE ION
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.6 |
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Grid | Model: Quantifoil R2/2 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number real images: 16721 / Average exposure time: 4.0 sec. / Average electron dose: 38.8 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 2.7 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.1 mm / Nominal defocus max: 2.8000000000000003 µm / Nominal defocus min: 0.8 µm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |