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Yorodumi- EMDB-13539: S. cerevisiae replisome-SCF(Dia2) complex bound to double-strande... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-13539 | ||||||||||||
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Title | S. cerevisiae replisome-SCF(Dia2) complex bound to double-stranded DNA (conformation II) | ||||||||||||
Map data | Composite cryo-EM density map for the budding yeast CMG-Csm3-Tof1-Mrc1-Ctf4-PolE-SCF(Dia2) complex on double-stranded DNA (conformation II), produced using the Phenix combine_focused_maps program | ||||||||||||
Sample |
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Function / homology | Function and homology information RAVE complex / Iron uptake and transport / CBF3 complex / regulation of transcription by galactose / regulation of sulfur amino acid metabolic process / cellular response to methylmercury / establishment of sister chromatid cohesion / vacuolar proton-transporting V-type ATPase complex assembly / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / DNA-templated DNA replication maintenance of fidelity ...RAVE complex / Iron uptake and transport / CBF3 complex / regulation of transcription by galactose / regulation of sulfur amino acid metabolic process / cellular response to methylmercury / establishment of sister chromatid cohesion / vacuolar proton-transporting V-type ATPase complex assembly / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / DNA-templated DNA replication maintenance of fidelity / gene conversion / maintenance of DNA repeat elements / septin ring assembly / Unwinding of DNA / invasive growth in response to glucose limitation / replication fork arrest / DNA replication initiation / Cul8-RING ubiquitin ligase complex / meiotic chromosome segregation / protein-containing complex disassembly / epsilon DNA polymerase complex / DNA strand elongation involved in mitotic DNA replication / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / nuclear DNA replication / MCM complex binding / GINS complex / mitotic DNA replication preinitiation complex assembly / premeiotic DNA replication / pre-replicative complex assembly involved in nuclear cell cycle DNA replication / nucleotide-excision repair, DNA gap filling / SUMO binding / mitotic DNA replication / regulation of exit from mitosis / DNA replication proofreading / Antigen processing: Ubiquitination & Proteasome degradation / Activation of the pre-replicative complex / CMG complex / single-stranded DNA 3'-5' DNA exonuclease activity / establishment of mitotic sister chromatid cohesion / DNA replication checkpoint signaling / nuclear pre-replicative complex / vacuolar acidification / kinetochore assembly / Activation of ATR in response to replication stress / MCM complex / regulation of metabolic process / DNA replication preinitiation complex / exit from mitosis / double-strand break repair via break-induced replication / mitotic DNA replication checkpoint signaling / replication fork protection complex / mitotic DNA replication initiation / single-stranded DNA helicase activity / positive regulation of glucose transmembrane transport / protein neddylation / mitotic intra-S DNA damage checkpoint signaling / Hydrolases; Acting on ester bonds; Exodeoxyribonucleases producing 5'-phosphomonoesters / regulation of DNA-templated DNA replication initiation / silent mating-type cassette heterochromatin formation / mitochondrial fusion / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / DNA strand elongation involved in DNA replication / SCF ubiquitin ligase complex / mitotic sister chromatid cohesion / leading strand elongation / nuclear chromosome / replication fork processing / DNA unwinding involved in DNA replication / nuclear replication fork / DNA replication origin binding / cullin family protein binding / regulation of DNA replication / subtelomeric heterochromatin formation / DNA replication initiation / endomembrane system / regulation of protein-containing complex assembly / error-prone translesion synthesis / negative regulation of cytoplasmic translation / heterochromatin formation / base-excision repair, gap-filling / regulation of mitotic cell cycle / DNA helicase activity / replication fork / meiotic cell cycle / helicase activity / G1/S transition of mitotic cell cycle / base-excision repair / DNA-templated DNA replication / kinetochore / nucleosome assembly / double-strand break repair via nonhomologous end joining / G2/M transition of mitotic cell cycle / double-strand break repair / single-stranded DNA binding / mitotic cell cycle / 4 iron, 4 sulfur cluster binding / ubiquitin-dependent protein catabolic process / double-stranded DNA binding Similarity search - Function | ||||||||||||
Biological species | DNA molecule (others) / Saccharomyces cerevisiae (brewer's yeast) / Synthetic construct (others) | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.2 Å | ||||||||||||
Authors | Jenkyn-Bedford M / Yeeles JTP / Deegan TD | ||||||||||||
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_13539.map.gz | 168 MB | EMDB map data format | |
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Header (meta data) | emd-13539-v30.xml emd-13539.xml | 49.7 KB 49.7 KB | Display Display | EMDB header |
Images | emd_13539.png | 95.4 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-13539 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13539 | HTTPS FTP |
-Related structure data
Related structure data | 7pmnMC 7ploC 7pmkC M: atomic model generated by this map C: citing same article (ref.) |
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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_13539.map.gz / Format: CCP4 / Size: 202.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Composite cryo-EM density map for the budding yeast CMG-Csm3-Tof1-Mrc1-Ctf4-PolE-SCF(Dia2) complex on double-stranded DNA (conformation II), produced using the Phenix combine_focused_maps program | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.06 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
+Entire : Budding yeast replisome on double-stranded DNA engaged with SCF(D...
+Supramolecule #1: Budding yeast replisome on double-stranded DNA engaged with SCF(D...
+Supramolecule #2: DNA
+Supramolecule #3: Replisome
+Macromolecule #1: DNA replication licensing factor MCM2
+Macromolecule #2: DNA replication licensing factor MCM3
+Macromolecule #3: DNA replication licensing factor MCM4
+Macromolecule #4: Minichromosome maintenance protein 5
+Macromolecule #5: DNA replication licensing factor MCM6
+Macromolecule #6: DNA replication licensing factor MCM7
+Macromolecule #7: DNA replication complex GINS protein PSF1
+Macromolecule #8: DNA replication complex GINS protein PSF2
+Macromolecule #9: DNA replication complex GINS protein PSF3
+Macromolecule #10: DNA replication complex GINS protein SLD5
+Macromolecule #11: Cell division control protein 45,Cell division control protein 45
+Macromolecule #12: DNA polymerase alpha-binding protein
+Macromolecule #15: Suppressor of kinetochore protein 1
+Macromolecule #16: Protein DIA2
+Macromolecule #17: DNA polymerase epsilon catalytic subunit A
+Macromolecule #18: DNA polymerase epsilon subunit B
+Macromolecule #19: Topoisomerase 1-associated factor 1
+Macromolecule #20: Chromosome segregation in meiosis protein 3
+Macromolecule #13: Leading strand template DNA
+Macromolecule #14: Lagging strand template DNA
+Macromolecule #21: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER
+Macromolecule #22: MAGNESIUM ION
+Macromolecule #23: ZINC 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 / Details: 15 mA |
Vitrification | Cryogen name: ETHANE / Details: Manual plunger. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.4 µm / Nominal magnification: 81000 |
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number real images: 12730 / Average exposure time: 4.0 sec. / Average electron dose: 38.8 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Particle selection | Number selected: 2160000 |
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CTF correction | Software - Name: RELION (ver. 3) |
Initial angle assignment | Type: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 3) |
Final angle assignment | Type: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 3) |
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3) Details: Composite map produced from individual cryo-EM density maps (resolutions 3.2 - 4.0 A) using Phenix combine_focused_maps. See publication for details. Number images used: 369254 |