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Yorodumi- PDB-8sok: Cryo-EM structure of human CST bound to POT1(ESDL)/TPP1 in the pr... -
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-Basic information
Entry | Database: PDB / ID: 8sok | |||||||||
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Title | Cryo-EM structure of human CST bound to POT1(ESDL)/TPP1 in the presence of telomeric ssDNA | |||||||||
Components |
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Keywords | DNA BINDING PROTEIN / telomere / shelterin / cst / complex | |||||||||
Function / homology | Function and homology information positive regulation of single-stranded telomeric DNA binding / positive regulation of DNA strand elongation / regulation of DNA helicase activity / positive regulation of DNA helicase activity / G-rich single-stranded DNA binding / telomere assembly / CST complex / segmentation / urogenital system development / positive regulation of helicase activity ...positive regulation of single-stranded telomeric DNA binding / positive regulation of DNA strand elongation / regulation of DNA helicase activity / positive regulation of DNA helicase activity / G-rich single-stranded DNA binding / telomere assembly / CST complex / segmentation / urogenital system development / positive regulation of helicase activity / 8-hydroxy-2'-deoxyguanosine DNA binding / regulation of double-strand break repair via nonhomologous end joining / telomeric D-loop binding / protection from non-homologous end joining at telomere / telomerase inhibitor activity / DEAD/H-box RNA helicase binding / establishment of protein localization to telomere / telomeric D-loop disassembly / telomere maintenance via telomere lengthening / shelterin complex / Telomere C-strand synthesis initiation / regulation of telomere maintenance via telomerase / telomere capping / Telomere C-strand (Lagging Strand) Synthesis / : / single-stranded telomeric DNA binding / positive regulation of telomere maintenance / nuclear telomere cap complex / Processive synthesis on the C-strand of the telomere / Polymerase switching on the C-strand of the telomere / Removal of the Flap Intermediate from the C-strand / G-rich strand telomeric DNA binding / telomerase holoenzyme complex / embryonic limb morphogenesis / bone marrow development / intermediate filament cytoskeleton / protein localization to chromosome, telomeric region / DNA duplex unwinding / hematopoietic stem cell proliferation / telomeric DNA binding / : / negative regulation of telomere maintenance via telomerase / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of MDA-5 activity / telomere maintenance via telomerase / Telomere Extension By Telomerase / replicative senescence / carbohydrate transmembrane transporter activity / DNA polymerase binding / Packaging Of Telomere Ends / spleen development / Recognition and association of DNA glycosylase with site containing an affected purine / Cleavage of the damaged purine / positive regulation of telomere maintenance via telomerase / regulation of G2/M transition of mitotic cell cycle / Recognition and association of DNA glycosylase with site containing an affected pyrimidine / Cleavage of the damaged pyrimidine / Inhibition of DNA recombination at telomere / telomere maintenance / Meiotic synapsis / picornain 2A / protein complex oligomerization / symbiont-mediated suppression of host mRNA export from nucleus / bioluminescence / symbiont genome entry into host cell via pore formation in plasma membrane / picornain 3C / monoatomic ion channel activity / thymus development / positive regulation of DNA replication / T=pseudo3 icosahedral viral capsid / skeletal system development / generation of precursor metabolites and energy / intracellular protein transport / host cell cytoplasmic vesicle membrane / cytoplasmic vesicle membrane / multicellular organism growth / DNA Damage/Telomere Stress Induced Senescence / endocytosis involved in viral entry into host cell / fibrillar center / positive regulation of fibroblast proliferation / nucleoside-triphosphate phosphatase / single-stranded DNA binding / outer membrane-bounded periplasmic space / chromosome, telomeric region / RNA helicase activity / nuclear body / induction by virus of host autophagy / RNA-directed RNA polymerase / viral RNA genome replication / cysteine-type endopeptidase activity / intracellular membrane-bounded organelle / RNA-dependent RNA polymerase activity / virus-mediated perturbation of host defense response / DNA-templated transcription / DNA damage response / protein-containing complex binding / virion attachment to host cell / structural molecule activity / ATP hydrolysis activity / proteolysis / RNA binding Similarity search - Function | |||||||||
Biological species | Escherichia coli O157:H7 (bacteria) Homo sapiens (human) Human enterovirus 71 synthetic construct (others) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.1 Å | |||||||||
Authors | Cai, S.W. | |||||||||
Funding support | United States, 2items
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Citation | Journal: Cell / Year: 2024 Title: POT1 recruits and regulates CST-Polα/primase at human telomeres. Authors: Sarah W Cai / Hiroyuki Takai / Arthur J Zaug / Teague C Dilgen / Thomas R Cech / Thomas Walz / Titia de Lange / Abstract: Telomere maintenance requires the extension of the G-rich telomeric repeat strand by telomerase and the fill-in synthesis of the C-rich strand by Polα/primase. At telomeres, Polα/primase is bound ...Telomere maintenance requires the extension of the G-rich telomeric repeat strand by telomerase and the fill-in synthesis of the C-rich strand by Polα/primase. At telomeres, Polα/primase is bound to Ctc1/Stn1/Ten1 (CST), a single-stranded DNA-binding complex. Like mutations in telomerase, mutations affecting CST-Polα/primase result in pathological telomere shortening and cause a telomere biology disorder, Coats plus (CP). We determined cryogenic electron microscopy structures of human CST bound to the shelterin heterodimer POT1/TPP1 that reveal how CST is recruited to telomeres by POT1. Our findings suggest that POT1 hinge phosphorylation is required for CST recruitment, and the complex is formed through conserved interactions involving several residues mutated in CP. Our structural and biochemical data suggest that phosphorylated POT1 holds CST-Polα/primase in an inactive, autoinhibited state until telomerase has extended the telomere ends. We propose that dephosphorylation of POT1 releases CST-Polα/primase into an active state that completes telomere replication through fill-in synthesis. #1: Journal: bioRxiv / Year: 2023 Title: POT1 recruits and regulates CST-Polα/Primase at human telomeres. Authors: Sarah W Cai / Hiroyuki Takai / Thomas Walz / Titia de Lange / Abstract: Telomere maintenance requires extension of the G-rich telomeric repeat strand by telomerase and fill-in synthesis of the C-rich strand by Polα/Primase. Telomeric Polα/Primase is bound to Ctc1-Stn1- ...Telomere maintenance requires extension of the G-rich telomeric repeat strand by telomerase and fill-in synthesis of the C-rich strand by Polα/Primase. Telomeric Polα/Primase is bound to Ctc1-Stn1-Ten1 (CST), a single-stranded DNA-binding complex. Like mutations in telomerase, mutations affecting CST-Polα/Primase result in pathological telomere shortening and cause a telomere biology disorder, Coats plus (CP). We determined cryogenic electron microscopy structures of human CST bound to the shelterin heterodimer POT1/TPP1 that reveal how CST is recruited to telomeres by POT1. Phosphorylation of POT1 is required for CST recruitment, and the complex is formed through conserved interactions involving several residues mutated in CP. Our structural and biochemical data suggest that phosphorylated POT1 holds CST-Polα/Primase in an inactive auto-inhibited state until telomerase has extended the telomere ends. We propose that dephosphorylation of POT1 releases CST-Polα/Primase into an active state that completes telomere replication through fill-in synthesis. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8sok.cif.gz | 828.7 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8sok.ent.gz | 655.5 KB | Display | PDB format |
PDBx/mmJSON format | 8sok.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8sok_validation.pdf.gz | 1.2 MB | Display | wwPDB validaton report |
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Full document | 8sok_full_validation.pdf.gz | 1.2 MB | Display | |
Data in XML | 8sok_validation.xml.gz | 75.4 KB | Display | |
Data in CIF | 8sok_validation.cif.gz | 113.9 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/so/8sok ftp://data.pdbj.org/pub/pdb/validation_reports/so/8sok | HTTPS FTP |
-Related structure data
Related structure data | 40660MC 8sojC M: map data used to model this data C: citing same article (ref.) |
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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
-CST complex subunit ... , 3 types, 3 molecules ABC
#1: Protein | Mass: 178300.609 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli O157:H7 (bacteria), (gene. exp.) Homo sapiens (human) Gene: malE, Z5632, ECs5017, CTC1, C17orf68 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P0AEY0, UniProt: Q2NKJ3 |
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#2: Protein | Mass: 42172.949 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: STN1, OBFC1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9H668 |
#3: Protein | Mass: 13872.013 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: TEN1, C17orf106 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q86WV5 |
-Protein , 2 types, 2 molecules DE
#4: Protein | Mass: 72938.219 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: POT1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9NUX5 |
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#5: Protein | Mass: 58064.863 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Human enterovirus 71, (gene. exp.) Homo sapiens (human) Gene: ACD, PIP1, PTOP, TINT1, TPP1 / Production host: Trichoplusia ni (cabbage looper) References: UniProt: B6F2F5, UniProt: Q96AP0, picornain 2A, nucleoside-triphosphate phosphatase, picornain 3C, RNA-directed RNA polymerase |
-DNA chain / Non-polymers , 2 types, 3 molecules F
#6: DNA chain | Mass: 5682.672 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
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#7: Chemical |
-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
Component | Name: ssDNA-bound human CST-POT1(ESDL)/TPP1 complex / Type: COMPLEX / Details: CST-POT1(ESDL)/TPP1 complex in the presence of DNA / Entity ID: #1-#6 / Source: RECOMBINANT |
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Molecular weight | Value: 0.37 MDa / Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Trichoplusia ni (cabbage looper) |
Buffer solution | pH: 7.5 |
Specimen | Conc.: 0.02 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: TFS KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm |
Image recording | Electron dose: 50.3 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
Software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||
3D reconstruction | Resolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 76359 / Symmetry type: POINT | ||||||||||||
Refinement | Cross valid method: NONE |