+ Open data
Open data
- Basic information
Basic information
| Entry | Database: PDB / ID: 6xty | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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| Title | CryoEM structure of human CMG bound to AND-1 (CMGA) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
|  Components | 
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|  Keywords | REPLICATION / CMG / Helicase / ATPase / Replisome | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Function / homology |  Function and homology information Switching of origins to a post-replicative state / Unwinding of DNA / DNA strand elongation involved in mitotic DNA replication / GINS complex / mitotic DNA replication preinitiation complex assembly / nuclear origin of replication recognition complex / Regulation of MITF-M-dependent genes involved in DNA replication, damage repair and senescence / alpha DNA polymerase:primase complex / mitotic DNA replication / DNA replication checkpoint signaling ...Switching of origins to a post-replicative state / Unwinding of DNA / DNA strand elongation involved in mitotic DNA replication / GINS complex / mitotic DNA replication preinitiation complex assembly / nuclear origin of replication recognition complex / Regulation of MITF-M-dependent genes involved in DNA replication, damage repair and senescence / alpha DNA polymerase:primase complex / mitotic DNA replication / DNA replication checkpoint signaling / regulation of phosphorylation / CMG complex / DNA replication preinitiation complex / MCM complex / double-strand break repair via break-induced replication / mitotic DNA replication initiation / regulation of DNA-templated DNA replication initiation / inner cell mass cell proliferation / DNA strand elongation involved in DNA replication / G1/S-Specific Transcription / nuclear replication fork / DNA replication origin binding / cochlea development / DNA replication initiation / Activation of the pre-replicative complex / Activation of ATR in response to replication stress / cellular response to interleukin-4 / DNA helicase activity / cellular response to epidermal growth factor stimulus / Assembly of the pre-replicative complex / helicase activity / DNA-templated DNA replication / multicellular organism growth / cellular response to xenobiotic stimulus / Orc1 removal from chromatin / cellular senescence / mitotic cell cycle / nucleosome assembly / single-stranded DNA binding / histone binding / DNA helicase / chromosome, telomeric region / DNA replication / cell population proliferation / cilium / ciliary basal body / DNA repair / intracellular membrane-bounded organelle / apoptotic process / DNA damage response / centrosome / chromatin binding / chromatin / perinuclear region of cytoplasm / enzyme binding / ATP hydrolysis activity / DNA binding / zinc ion binding / nucleoplasm / ATP binding / identical protein binding / nucleus / membrane / cytoplasm / cytosol Similarity search - Function | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Biological species |  Homo sapiens (human) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 6.77 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
|  Authors | Rzechorzek, N.J. / Pellegrini, L. / Chirgadze, D.Y. / Hardwick, S.W. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Funding support |  United Kingdom, 1items 
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|  Citation |  Journal: Nucleic Acids Res / Year: 2020 Title: CryoEM structures of human CMG-ATPγS-DNA and CMG-AND-1 complexes. Authors: Neil J Rzechorzek / Steven W Hardwick / Vincentius A Jatikusumo / Dimitri Y Chirgadze / Luca Pellegrini /  Abstract: DNA unwinding in eukaryotic replication is performed by the Cdc45-MCM-GINS (CMG) helicase. Although the CMG architecture has been elucidated, its mechanism of DNA unwinding and replisome interactions ...DNA unwinding in eukaryotic replication is performed by the Cdc45-MCM-GINS (CMG) helicase. Although the CMG architecture has been elucidated, its mechanism of DNA unwinding and replisome interactions remain poorly understood. Here we report the cryoEM structure at 3.3 Å of human CMG bound to fork DNA and the ATP-analogue ATPγS. Eleven nucleotides of single-stranded (ss) DNA are bound within the C-tier of MCM2-7 AAA+ ATPase domains. All MCM subunits contact DNA, from MCM2 at the 5'-end to MCM5 at the 3'-end of the DNA spiral, but only MCM6, 4, 7 and 3 make a full set of interactions. DNA binding correlates with nucleotide occupancy: five MCM subunits are bound to either ATPγS or ADP, whereas the apo MCM2-5 interface remains open. We further report the cryoEM structure of human CMG bound to the replisome hub AND-1 (CMGA). The AND-1 trimer uses one β-propeller domain of its trimerisation region to dock onto the side of the helicase assembly formed by Cdc45 and GINS. In the resulting CMGA architecture, the AND-1 trimer is closely positioned to the fork DNA while its CIP (Ctf4-interacting peptide)-binding helical domains remain available to recruit partner proteins. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| History | 
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- Structure visualization
Structure visualization
| Movie | 
 
  Movie viewer | 
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| Structure viewer | Molecule:  Molmil  Jmol/JSmol | 
- Downloads & links
Downloads & links
- Download
Download
| PDBx/mmCIF format |  6xty.cif.gz | 1.1 MB | Display |  PDBx/mmCIF format | 
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| PDB format |  pdb6xty.ent.gz | 913.9 KB | Display |  PDB format | 
| PDBx/mmJSON format |  6xty.json.gz | Tree view |  PDBx/mmJSON format | |
| Others |  Other downloads | 
-Validation report
| Summary document |  6xty_validation.pdf.gz | 1.7 MB | Display |  wwPDB validaton report | 
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| Full document |  6xty_full_validation.pdf.gz | 1.8 MB | Display | |
| Data in XML |  6xty_validation.xml.gz | 176.2 KB | Display | |
| Data in CIF |  6xty_validation.cif.gz | 267.6 KB | Display | |
| Arichive directory |  https://data.pdbj.org/pub/pdb/validation_reports/xt/6xty  ftp://data.pdbj.org/pub/pdb/validation_reports/xt/6xty | HTTPS FTP | 
-Related structure data
| Related structure data |  10621MC  6xtxC C: citing same article ( M: map data used to model this data | 
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| Similar structure data | |
| EM raw data |  EMPIAR-10472 (Title: CryoEM structure of human CMG bound to AND-1 (CMGA) / Data size: 1.3 TB Data #1: Unaligned multiframe micrographs of human CMG bound to AND-1 (CMGA) [micrographs - multiframe]) | 
- Links
Links
- Assembly
Assembly
| Deposited unit |  
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| 1 | 
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- Components
Components
-DNA replication licensing factor  ... , 6 types, 6 molecules 234567     
| #1: Protein | Mass: 102034.102 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: MCM2, BM28, CCNL1, CDCL1, KIAA0030 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: P49736, DNA helicase | 
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| #2: Protein | Mass: 96043.320 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: MCM3 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: P25205, DNA helicase | 
| #3: Protein | Mass: 96684.852 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: MCM4, CDC21 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: P33991, DNA helicase | 
| #4: Protein | Mass: 82406.633 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: MCM5, CDC46 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: P33992, DNA helicase | 
| #5: Protein | Mass: 93010.273 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: MCM6 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: Q14566, DNA helicase | 
| #6: Protein | Mass: 81411.875 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: MCM7, CDC47, MCM2 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: P33993, DNA helicase | 
-DNA replication complex GINS protein  ... , 4 types, 4 molecules ABCD   
| #7: Protein | Mass: 23022.469 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: GINS1, KIAA0186, PSF1 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: Q14691 | 
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| #8: Protein | Mass: 21453.713 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: GINS2, PSF2, CGI-122, DC5, HSPC037 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: Q9Y248 | 
| #9: Protein | Mass: 24562.611 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: GINS3, PSF3 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: Q9BRX5 | 
| #10: Protein | Mass: 26081.873 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: GINS4, SLD5 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: Q9BRT9 | 
-Protein , 2 types, 4 molecules EFGH   
| #11: Protein | Mass: 65650.555 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: CDC45, CDC45L, CDC45L2, UNQ374/PRO710 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: O75419 | 
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| #12: Protein | Mass: 130484.750 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.)  Homo sapiens (human) / Gene: WDHD1, AND1 / Cell line (production host): HEK293F / Production host:  Homo sapiens (human) / References: UniProt: O75717 | 
-Non-polymers , 1 types, 5 molecules 
| #13: Chemical | ChemComp-ZN / | 
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-Details
| Has ligand of interest | N | 
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| Has protein modification | N | 
-Experimental details
-Experiment
| Experiment | Method: ELECTRON MICROSCOPY | 
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| EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction | 
- Sample preparation
Sample preparation
| Component | 
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| Source (natural) | 
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| Source (recombinant) | 
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| Buffer solution | pH: 7.5 | ||||||||||||||||||||||||
| Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||
| Vitrification | Cryogen name: ETHANE | 
- Electron microscopy imaging
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 | Electron dose: 54.3 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k) | 
- Processing
Processing
| Software | Name: PHENIX / Version: 1.16_3549: / Classification: refinement | 
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| EM software | Name: PHENIX / Category: model refinement | 
| CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | 
| Symmetry | Point symmetry: C1 (asymmetric) | 
| 3D reconstruction | Resolution: 6.77 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 15393 / Symmetry type: POINT | 
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