+Open data
-Basic information
Entry | Database: PDB / ID: 7jk3 | ||||||
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Title | Structure of Drosophila ORC bound to GC-rich DNA and Cdc6 | ||||||
Components |
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Keywords | REPLICATION/DNA / origin recognition complex / DNA replication initiation / REPLICATION / REPLICATION-DNA complex | ||||||
Function / homology | Function and homology information alpha-heterochromatin / larval feeding behavior / CDC6 association with the ORC:origin complex / septin cytoskeleton / CDK-mediated phosphorylation and removal of Cdc6 / septin cytoskeleton organization / Activation of ATR in response to replication stress / Activation of the pre-replicative complex / eggshell chorion gene amplification / Orc1 removal from chromatin ...alpha-heterochromatin / larval feeding behavior / CDC6 association with the ORC:origin complex / septin cytoskeleton / CDK-mediated phosphorylation and removal of Cdc6 / septin cytoskeleton organization / Activation of ATR in response to replication stress / Activation of the pre-replicative complex / eggshell chorion gene amplification / Orc1 removal from chromatin / origin recognition complex / DNA amplification / positive regulation of border follicle cell migration / Assembly of the ORC complex at the origin of replication / nuclear origin of replication recognition complex / olfactory learning / nuclear pre-replicative complex / DNA replication preinitiation complex / mitotic DNA replication checkpoint signaling / mitotic chromosome condensation / chromosome condensation / DNA replication origin binding / DNA replication initiation / heterochromatin / ribonucleoprotein complex binding / nuclear pore / GTPase activator activity / mitotic spindle organization / learning / DNA-templated DNA replication / mitotic cell cycle / DNA replication / learning or memory / cell division / chromatin binding / protein homodimerization activity / ATP hydrolysis activity / DNA binding / ATP binding / nucleus / metal ion binding / plasma membrane Similarity search - Function | ||||||
Biological species | Drosophila melanogaster (fruit fly) synthetic construct (others) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||
Authors | Schmidt, J.M. / Bleichert, F. | ||||||
Funding support | Switzerland, 1items
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Citation | Journal: Nat Commun / Year: 2020 Title: Structural mechanism for replication origin binding and remodeling by a metazoan origin recognition complex and its co-loader Cdc6. Authors: Jan Marten Schmidt / Franziska Bleichert / Abstract: Eukaryotic DNA replication initiation relies on the origin recognition complex (ORC), a DNA-binding ATPase that loads the Mcm2-7 replicative helicase onto replication origins. Here, we report cryo- ...Eukaryotic DNA replication initiation relies on the origin recognition complex (ORC), a DNA-binding ATPase that loads the Mcm2-7 replicative helicase onto replication origins. Here, we report cryo-electron microscopy (cryo-EM) structures of DNA-bound Drosophila ORC with and without the co-loader Cdc6. These structures reveal that Orc1 and Orc4 constitute the primary DNA binding site in the ORC ring and cooperate with the winged-helix domains to stabilize DNA bending. A loop region near the catalytic Walker B motif of Orc1 directly contacts DNA, allosterically coupling DNA binding to ORC's ATPase site. Correlating structural and biochemical data show that DNA sequence modulates DNA binding and remodeling by ORC, and that DNA bending promotes Mcm2-7 loading in vitro. Together, these findings explain the distinct DNA sequence-dependencies of metazoan and S. cerevisiae initiators in origin recognition and support a model in which DNA geometry and bendability contribute to Mcm2-7 loading site selection in metazoans. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7jk3.cif.gz | 514.9 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7jk3.ent.gz | 396.4 KB | Display | PDB format |
PDBx/mmJSON format | 7jk3.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7jk3_validation.pdf.gz | 1.6 MB | Display | wwPDB validaton report |
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Full document | 7jk3_full_validation.pdf.gz | 1.6 MB | Display | |
Data in XML | 7jk3_validation.xml.gz | 77.9 KB | Display | |
Data in CIF | 7jk3_validation.cif.gz | 119 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/jk/7jk3 ftp://data.pdbj.org/pub/pdb/validation_reports/jk/7jk3 | HTTPS FTP |
-Related structure data
Related structure data | 22360MC 7jgrC 7jgsC 7jk2C 7jk4C 7jk5C 7jk6C C: citing same article (ref.) M: map data used to model this data |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Origin recognition complex subunit ... , 6 types, 6 molecules BCDEFA
#1: Protein | Mass: 69086.445 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Drosophila melanogaster (fruit fly) / Gene: Orc2, CG3041 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q24168 |
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#2: Protein | Mass: 82364.523 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Drosophila melanogaster (fruit fly) Gene: Orc3, CG34315-RB, Dmel\CG4088, DmORC3, DmOrc3, l(2)49Fd, l(2)vr6, LAT, Lat, lat, lat-RA, latheo, ORC, ORC3, orc3, vr-6, vr6, CG4088, Dmel_CG4088 Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q7K2L1 |
#3: Protein | Mass: 52277.109 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Drosophila melanogaster (fruit fly) Gene: Orc4, Dmel\CG2917, DmORC4, dmOrc4, ORC, ORC4, orc4, rDmORC, CG2917, Dmel_CG2917 Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9W102 |
#4: Protein | Mass: 52175.066 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Drosophila melanogaster (fruit fly) / Gene: Orc5, CG7833 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q24169 |
#5: Protein | Mass: 29284.129 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Drosophila melanogaster (fruit fly) / Gene: Orc6, CG1584 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9Y1B2 |
#6: Protein | Mass: 54485.633 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Drosophila melanogaster (fruit fly) / Gene: Orc1, CG10667 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: O16810 |
-Protein , 1 types, 1 molecules G
#7: Protein | Mass: 48049.465 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Drosophila melanogaster (fruit fly) Gene: Cdc6, CDC6, cdc6, DmCdc6, Dmel\CG5971, CG5971, Dmel_CG5971 Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9VSM9 |
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-DNA chain , 2 types, 2 molecules HI
#8: DNA chain | Mass: 18679.869 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
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#9: DNA chain | Mass: 18315.686 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
-Non-polymers , 2 types, 8 molecules
#10: Chemical | ChemComp-ATP / #11: Chemical | ChemComp-MG / |
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-Details
Has ligand of interest | N |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Drosophila ORC bound to GC-rich DNA and Cdc6 / Type: COMPLEX / Entity ID: #1-#9 / Source: RECOMBINANT | ||||||||||||||||||||||||
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Molecular weight | Experimental value: NO | ||||||||||||||||||||||||
Source (natural) | Organism: Drosophila melanogaster (fruit fly) | ||||||||||||||||||||||||
Source (recombinant) | Organism: Trichoplusia ni (cabbage looper) | ||||||||||||||||||||||||
Buffer solution | pH: 7.6 | ||||||||||||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: Recombinantly expressed and purified Drosophila ORC and Cdc6 were mixed with DNA and the DNA-bound ternary complex purified by size exclusion chromatography. | ||||||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil, UltrAuFoil, R1.2/1.3 | ||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company | ||||||||||||||||||
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Microscopy | Model: FEI TITAN KRIOS | ||||||||||||||||||
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM | ||||||||||||||||||
Electron lens | Mode: BRIGHT FIELD | ||||||||||||||||||
Image recording |
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-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 174891 / Symmetry type: POINT |