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Yorodumi- PDB-8b8t: Open conformation of the complex of DNA ligase I on PCNA and DNA ... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8b8t | ||||||
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Title | Open conformation of the complex of DNA ligase I on PCNA and DNA in the presence of ATP | ||||||
Components |
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Keywords | REPLICATION / DNA / Complex / Ligase / PCNA / Ligation / Okazaki fragment maturation | ||||||
Function / homology | Function and homology information Okazaki fragment processing involved in mitotic DNA replication / Regulation of MITF-M-dependent genes involved in DNA damage repair and senescence / DNA ligase activity / positive regulation of deoxyribonuclease activity / dinucleotide insertion or deletion binding / DNA ligase (ATP) / PCNA-p21 complex / DNA ligase (ATP) activity / mitotic telomere maintenance via semi-conservative replication / purine-specific mismatch base pair DNA N-glycosylase activity ...Okazaki fragment processing involved in mitotic DNA replication / Regulation of MITF-M-dependent genes involved in DNA damage repair and senescence / DNA ligase activity / positive regulation of deoxyribonuclease activity / dinucleotide insertion or deletion binding / DNA ligase (ATP) / PCNA-p21 complex / DNA ligase (ATP) activity / mitotic telomere maintenance via semi-conservative replication / purine-specific mismatch base pair DNA N-glycosylase activity / nuclear lamina / MutLalpha complex binding / positive regulation of DNA-directed DNA polymerase activity / Polymerase switching / Telomere C-strand (Lagging Strand) Synthesis / Processive synthesis on the lagging strand / DNA ligation / PCNA complex / Removal of the Flap Intermediate / Processive synthesis on the C-strand of the telomere / Mismatch repair (MMR) directed by MSH2:MSH3 (MutSbeta) / Polymerase switching on the C-strand of the telomere / Mismatch repair (MMR) directed by MSH2:MSH6 (MutSalpha) / Transcription of E2F targets under negative control by DREAM complex / Removal of the Flap Intermediate from the C-strand / lagging strand elongation / replisome / response to L-glutamate / histone acetyltransferase binding / DNA biosynthetic process / DNA polymerase processivity factor activity / positive regulation of DNA replication / G1/S-Specific Transcription / replication fork processing / response to dexamethasone / leading strand elongation / Early Phase of HIV Life Cycle / nuclear replication fork / cyclin-dependent protein kinase holoenzyme complex / SUMOylation of DNA replication proteins / estrous cycle / POLB-Dependent Long Patch Base Excision Repair / epithelial cell differentiation / PCNA-Dependent Long Patch Base Excision Repair / anatomical structure morphogenesis / positive regulation of DNA repair / mismatch repair / male germ cell nucleus / translesion synthesis / response to cadmium ion / DNA polymerase binding / liver regeneration / Translesion synthesis by REV1 / Translesion synthesis by POLK / base-excision repair, gap-filling / Translesion synthesis by POLI / Gap-filling DNA repair synthesis and ligation in GG-NER / TP53 Regulates Transcription of Genes Involved in G2 Cell Cycle Arrest / replication fork / nuclear estrogen receptor binding / Recognition of DNA damage by PCNA-containing replication complex / Termination of translesion DNA synthesis / Translesion Synthesis by POLH / base-excision repair / HDR through Homologous Recombination (HRR) / Dual Incision in GG-NER / receptor tyrosine kinase binding / cellular response to hydrogen peroxide / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / cellular response to xenobiotic stimulus / cellular response to UV / E3 ubiquitin ligases ubiquitinate target proteins / response to estradiol / heart development / chromosome, telomeric region / DNA recombination / damaged DNA binding / nuclear body / cell division / intracellular membrane-bounded organelle / DNA repair / chromatin binding / centrosome / chromatin / protein-containing complex binding / enzyme binding / negative regulation of transcription by RNA polymerase II / mitochondrion / DNA binding / extracellular exosome / nucleoplasm / ATP binding / identical protein binding / nucleus / metal ion binding Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.2 Å | ||||||
Authors | Blair, K. / Tehseen, M. / Raducanu, V.S. / Shahid, T. / Lancey, C. / Cruehet, R. / Hamdan, S. / De Biasio, A. | ||||||
Funding support | United Kingdom, 1items
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Citation | Journal: Nat Commun / Year: 2022 Title: Mechanism of human Lig1 regulation by PCNA in Okazaki fragment sealing. Authors: Kerry Blair / Muhammad Tehseen / Vlad-Stefan Raducanu / Taha Shahid / Claudia Lancey / Fahad Rashid / Ramon Crehuet / Samir M Hamdan / Alfredo De Biasio / Abstract: During lagging strand synthesis, DNA Ligase 1 (Lig1) cooperates with the sliding clamp PCNA to seal the nicks between Okazaki fragments generated by Pol δ and Flap endonuclease 1 (FEN1). We present ...During lagging strand synthesis, DNA Ligase 1 (Lig1) cooperates with the sliding clamp PCNA to seal the nicks between Okazaki fragments generated by Pol δ and Flap endonuclease 1 (FEN1). We present several cryo-EM structures combined with functional assays, showing that human Lig1 recruits PCNA to nicked DNA using two PCNA-interacting motifs (PIPs) located at its disordered N-terminus (PIP) and DNA binding domain (PIP). Once Lig1 and PCNA assemble as two-stack rings encircling DNA, PIP is released from PCNA and only PIP is required for ligation to facilitate the substrate handoff from FEN1. Consistently, we observed that PCNA forms a defined complex with FEN1 and nicked DNA, and it recruits Lig1 to an unoccupied monomer creating a toolbelt that drives the transfer of DNA to Lig1. Collectively, our results provide a structural model on how PCNA regulates FEN1 and Lig1 during Okazaki fragments maturation. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8b8t.cif.gz | 175 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8b8t.ent.gz | 134.9 KB | Display | PDB format |
PDBx/mmJSON format | 8b8t.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8b8t_validation.pdf.gz | 1.1 MB | Display | wwPDB validaton report |
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Full document | 8b8t_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 8b8t_validation.xml.gz | 36.9 KB | Display | |
Data in CIF | 8b8t_validation.cif.gz | 57.5 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/b8/8b8t ftp://data.pdbj.org/pub/pdb/validation_reports/b8/8b8t | HTTPS FTP |
-Related structure data
Related structure data | 15921MC 7qnzC 7qo1C C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 29720.342 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LIG1 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P18858, DNA ligase (ATP) |
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#2: Protein | Mass: 28441.504 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: PCNA / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P12004 |
-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: complex of DNA ligase I on PCNA and DNA in the presence of ATP Type: COMPLEX / Entity ID: all / Source: RECOMBINANT | ||||||||||||||||||||||||||||||
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Source (natural) | Organism: Homo sapiens (human) | ||||||||||||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) | ||||||||||||||||||||||||||||||
Buffer solution | pH: 7.5 | ||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-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 / Nominal magnification: 105000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: ZEMLIN TABLEAU |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 77 K / Temperature (min): 77 K |
Image recording | Average exposure time: 2 sec. / Electron dose: 18 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
Image scans | Width: 5760 / Height: 4092 |
-Processing
Software | Name: PHENIX / Version: 1.19.2_4158: / Classification: refinement | ||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 107550 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT | ||||||||||||||||||||||||
Refine LS restraints |
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