[English] 日本語
Yorodumi- PDB-8w0a: Human DNA polymerase theta helicase domain in complex with ssDNA,... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8w0a | ||||||
---|---|---|---|---|---|---|---|
Title | Human DNA polymerase theta helicase domain in complex with ssDNA, dimer form | ||||||
Components |
| ||||||
Keywords | TRANSFERASE/DNA / DNA repair / helicase / ATPase / TRANSFERASE-DNA complex | ||||||
Function / homology | Function and homology information single-stranded DNA endodeoxyribonuclease activity / HDR through MMEJ (alt-NHEJ) / double-strand break repair via alternative nonhomologous end joining / single-stranded DNA helicase activity / replication fork processing / site of DNA damage / negative regulation of double-strand break repair via homologous recombination / 5'-deoxyribose-5-phosphate lyase activity / error-prone translesion synthesis / somatic hypermutation of immunoglobulin genes ...single-stranded DNA endodeoxyribonuclease activity / HDR through MMEJ (alt-NHEJ) / double-strand break repair via alternative nonhomologous end joining / single-stranded DNA helicase activity / replication fork processing / site of DNA damage / negative regulation of double-strand break repair via homologous recombination / 5'-deoxyribose-5-phosphate lyase activity / error-prone translesion synthesis / somatic hypermutation of immunoglobulin genes / DNA helicase activity / base-excision repair / protein homooligomerization / RNA-directed DNA polymerase / RNA-directed DNA polymerase activity / double-strand break repair / site of double-strand break / DNA helicase / damaged DNA binding / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / DNA repair / DNA damage response / chromatin binding / Golgi apparatus / magnesium ion binding / ATP hydrolysis activity / nucleoplasm / ATP binding / identical protein binding / cytosol Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å | ||||||
Authors | Ito, F. / Li, Z. / Chen, X.S. | ||||||
Funding support | United States, 1items
| ||||||
Citation | Journal: bioRxiv / Year: 2024 Title: Structural Basis for Polθ-Helicase DNA Binding and Microhomology-Mediated End-Joining. Authors: Fumiaki Ito / Ziyuan Li / Leonid Minakhin / Htet A Khant / Richard T Pomerantz / Xiaojiang S Chen / Abstract: DNA double-strand breaks (DSBs) present a critical threat to genomic integrity, often precipitating genomic instability and oncogenesis. Repair of DSBs predominantly occurs through homologous ...DNA double-strand breaks (DSBs) present a critical threat to genomic integrity, often precipitating genomic instability and oncogenesis. Repair of DSBs predominantly occurs through homologous recombination (HR) and non-homologous end joining (NHEJ). In HR-deficient cells, DNA polymerase theta (Polθ) becomes critical for DSB repair via microhomology-mediated end joining (MMEJ), also termed theta-mediated end joining (TMEJ). Thus, Polθ is synthetically lethal with BRCA1/2 and other HR factors, underscoring its potential as a therapeutic target in HR-deficient cancers. However, the molecular mechanisms governing Polθ-mediated MMEJ remain poorly understood. Here we present a series of cryo-electron microscopy structures of the Polθ helicase domain (Polθ-hel) in complex with DNA containing 3'-overhang. The structures reveal the sequential conformations adopted by Polθ-hel during the critical phases of DNA binding, microhomology searching, and microhomology annealing. The stepwise conformational changes within the Polθ-hel subdomains and its functional dimeric state are pivotal for aligning the 3'-overhangs, facilitating the microhomology search and subsequent annealing necessary for DSB repair via MMEJ. Our findings illustrate the essential molecular switches within Polθ-hel that orchestrate the MMEJ process in DSB repair, laying the groundwork for the development of targeted therapies against the Polθ-hel. | ||||||
History |
|
-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
---|
-Downloads & links
-Download
PDBx/mmCIF format | 8w0a.cif.gz | 267.1 KB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb8w0a.ent.gz | 208.3 KB | Display | PDB format |
PDBx/mmJSON format | 8w0a.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8w0a_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
---|---|---|---|---|
Full document | 8w0a_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 8w0a_validation.xml.gz | 54.4 KB | Display | |
Data in CIF | 8w0a_validation.cif.gz | 78.9 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/w0/8w0a ftp://data.pdbj.org/pub/pdb/validation_reports/w0/8w0a | HTTPS FTP |
-Related structure data
Related structure data | 43706MC 9asjC 9askC 9aslC 9c5qC 40760 40761 M: map data used to model this data C: citing same article (ref.) |
---|---|
Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
|
---|---|
1 |
|
-Components
#1: Protein | Mass: 99802.539 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: POLQ, POLH / Production host: Escherichia coli (E. coli) / References: UniProt: O75417, DNA-directed DNA polymerase #2: DNA chain | Mass: 13480.646 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Human DNA polymerase theta helicase domain in complex with ssDNA Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES |
---|---|
Molecular weight | Value: 0.253 MDa / Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Escherichia coli (E. coli) |
Buffer solution | pH: 8 |
Specimen | Conc.: 0.8 mg/ml / 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 |
---|---|
Microscopy | Model: TFS 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: 3000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm |
Image recording | Average exposure time: 3.5 sec. / Electron dose: 65 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 7228 |
-Processing
EM software | Name: EPU / Category: image acquisition |
---|---|
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION |
Particle selection | Num. of particles selected: 2654648 |
Symmetry | Point symmetry: C2 (2 fold cyclic) |
3D reconstruction | Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 99902 / Symmetry type: POINT |