+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 / Switching of origins to a post-replicative state / detection of abiotic stimulus / replication fork arrest / regulation of nuclear cell cycle DNA replication / Unwinding of DNA / DNA replication initiation / epsilon DNA polymerase complex / cell cycle phase transition ...cellular response to bleomycin / DNA secondary structure binding / Switching of origins to a post-replicative state / detection of abiotic stimulus / replication fork arrest / regulation of nuclear cell cycle DNA replication / Unwinding of DNA / DNA replication initiation / epsilon DNA polymerase complex / cell cycle phase transition / cellular response to cisplatin / GINS complex / DNA strand elongation involved in mitotic DNA replication / mitotic DNA replication preinitiation complex assembly / nuclear origin of replication recognition complex / cellular response to hydroxyurea / alpha DNA polymerase:primase complex / mitotic DNA replication / cullin-RING-type E3 NEDD8 transferase / anaphase-promoting complex binding / cellular response to chemical stress / NEDD8 transferase activity / CMG complex / cullin-RING ubiquitin ligase complex / DNA replication checkpoint signaling / single-stranded DNA 3'-5' DNA exonuclease activity / single-stranded 3'-5' DNA helicase activity / entrainment of circadian clock / Cul7-RING ubiquitin ligase complex / ubiquitin-dependent protein catabolic process via the C-end degron rule pathway / regulation of phosphorylation / MCM complex / DNA replication preinitiation complex / Loss of Function of FBXW7 in Cancer and NOTCH1 Signaling / target-directed miRNA degradation / elongin complex / VCB complex / replication fork protection complex / positive regulation of protein autoubiquitination / mitotic DNA replication checkpoint signaling / mitotic DNA replication initiation / protein neddylation / double-strand break repair via break-induced replication / NEDD8 ligase activity / mitotic intra-S DNA damage checkpoint signaling / positive regulation of double-strand break repair / Cul5-RING ubiquitin ligase complex / negative regulation of response to oxidative stress / Hydrolases; Acting on ester bonds; Exodeoxyribonucleases producing 5'-phosphomonoesters / regulation of DNA-templated DNA replication initiation / ubiquitin-ubiquitin ligase activity / Cul4A-RING E3 ubiquitin ligase complex / single-stranded DNA helicase activity / SCF ubiquitin ligase complex / inner cell mass cell proliferation / Cul2-RING ubiquitin ligase complex / negative regulation of type I interferon production / nucleotide-excision repair, DNA gap filling / DNA replication proofreading / Cul4B-RING E3 ubiquitin ligase complex / ubiquitin ligase complex scaffold activity / DNA strand elongation involved in DNA replication / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / Cul3-RING ubiquitin ligase complex / branching morphogenesis of an epithelial tube / DNA synthesis involved in DNA repair / cochlea development / G1/S-Specific Transcription / activation of protein kinase activity / leading strand elongation / replication fork processing / Prolactin receptor signaling / Apoptotic cleavage of cellular proteins / DNA unwinding involved in DNA replication / nuclear replication fork / protein monoubiquitination / mitotic G2 DNA damage checkpoint signaling / 3'-5' DNA helicase activity / 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 / Activation of the pre-replicative complex / PCNA-Dependent Long Patch Base Excision Repair / DNA replication initiation / Tat-mediated elongation of the HIV-1 transcript / Formation of HIV-1 elongation complex containing HIV-1 Tat / cellular response to interleukin-4 / embryonic organ development / Nuclear events stimulated by ALK signaling in cancer / protein K48-linked ubiquitination / Formation of HIV elongation complex in the absence of HIV Tat / error-prone translesion synthesis / Activation of ATR in response to replication stress / RNA Polymerase II Transcription Elongation / Formation of RNA Pol II elongation complex / response to UV 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 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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 |