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Yorodumi- EMDB-42384: Cryo-EM map of the human CTF18-RFC-PCNA binary complex in the fou... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-42384 | |||||||||
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Title | Cryo-EM map of the human CTF18-RFC-PCNA binary complex in the four-subunit binding state (state 3) | |||||||||
Map data | Primary sharp EM map | |||||||||
Sample |
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Keywords | DNA clamp loader complex / REPLICATION-DNA complex | |||||||||
Function / homology | Function and homology information positive regulation of deoxyribonuclease activity / dinucleotide insertion or deletion binding / PCNA-p21 complex / response to organophosphorus / Ctf18 RFC-like complex / DNA clamp loader activity / DNA replication factor C complex / Elg1 RFC-like complex / mitotic telomere maintenance via semi-conservative replication / purine-specific mismatch base pair DNA N-glycosylase activity ...positive regulation of deoxyribonuclease activity / dinucleotide insertion or deletion binding / PCNA-p21 complex / response to organophosphorus / Ctf18 RFC-like complex / DNA clamp loader activity / DNA replication factor C complex / Elg1 RFC-like complex / 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 / 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 / replisome / Activation of ATR in response to replication stress / DNA duplex unwinding / response to L-glutamate / HDR through Single Strand Annealing (SSA) / Impaired BRCA2 binding to RAD51 / DNA strand elongation involved in DNA replication / histone acetyltransferase binding / DNA synthesis involved in DNA repair / DNA polymerase processivity factor activity / positive regulation of DNA replication / G1/S-Specific Transcription / replication fork processing / response to dexamethasone / leading strand elongation / nuclear replication fork / cyclin-dependent protein kinase holoenzyme complex / Presynaptic phase of homologous DNA pairing and strand exchange / SUMOylation of DNA replication proteins / estrous cycle / epithelial cell differentiation / PCNA-Dependent Long Patch Base Excision Repair / positive regulation of DNA repair / ATP-dependent activity, acting on DNA / 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 / G2/M DNA damage checkpoint / Translesion Synthesis by POLH / HDR through Homologous Recombination (HRR) / Dual Incision in GG-NER / DNA-templated DNA replication / 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 / Processing of DNA double-strand break ends / E3 ubiquitin ligases ubiquitinate target proteins / response to estradiol / heart development / chromosome, telomeric region / DNA replication / Regulation of TP53 Activity through Phosphorylation / damaged DNA binding / nuclear body / DNA repair / chromatin binding / centrosome / chromatin / protein-containing complex binding / enzyme binding / negative regulation of transcription by RNA polymerase II / ATP hydrolysis activity / DNA binding / extracellular exosome / nucleoplasm / ATP binding / identical protein binding / membrane / nucleus / cytosol Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.16 Å | |||||||||
Authors | Wang F / He Q / Li H | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2024 Title: Cryo-EM reveals a nearly complete PCNA loading process and unique features of the human alternative clamp loader CTF18-RFC. Authors: Qing He / Feng Wang / Michael E O'Donnell / Huilin Li / Abstract: The DNA sliding clamp PCNA is a multipurpose platform for DNA polymerases and many other proteins involved in DNA metabolism. The topologically closed PCNA ring needs to be cracked open and loaded ...The DNA sliding clamp PCNA is a multipurpose platform for DNA polymerases and many other proteins involved in DNA metabolism. The topologically closed PCNA ring needs to be cracked open and loaded onto DNA by a clamp loader, e.g., the well-studied pentameric ATPase complex RFC (RFC1-5). The CTF18-RFC complex is an alternative clamp loader found recently to bind the leading strand DNA polymerase ε and load PCNA onto leading strand DNA, but its structure and the loading mechanism have been unknown. By cryo-EM analysis of in vitro assembled human CTF18-RFC-DNA-PCNA complex, we have captured seven loading intermediates, revealing a detailed PCNA loading mechanism onto a 3'-ss/dsDNA junction by CTF18-RFC. Interestingly, the alternative loader has evolved a highly mobile CTF18 AAA+ module likely to lower the loading activity, perhaps to avoid competition with the RFC and to limit its role to leading strand clamp loading. To compensate for the lost stability due to the mobile AAA+ module, CTF18 has evolved a unique β-hairpin motif that reaches across RFC2 to interact with RFC5, thereby stabilizing the pentameric complex. Further, we found that CTF18 also contains a separation pin to locally melt DNA from the 3'-end of the primer; this ensures its ability to load PCNA to any 3'-ss/dsDNA junction, facilitated by the binding energy of the E-plug to the major groove. Our study reveals unique structural features of the human CTF18-RFC and contributes to a broader understanding of PCNA loading by the alternative clamp loaders. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_42384.map.gz | 118 MB | EMDB map data format | |
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Header (meta data) | emd-42384-v30.xml emd-42384.xml | 27.4 KB 27.4 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_42384_fsc.xml | 10.6 KB | Display | FSC data file |
Images | emd_42384.png | 52.5 KB | ||
Filedesc metadata | emd-42384.cif.gz | 7.9 KB | ||
Others | emd_42384_additional_1.map.gz emd_42384_additional_2.map.gz emd_42384_half_map_1.map.gz emd_42384_half_map_2.map.gz | 62 MB 111 MB 116.2 MB 116.2 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-42384 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-42384 | HTTPS FTP |
-Related structure data
Related structure data | 8umuMC 8umtC 8umvC 8umwC 8umyC 8un0C 8unjC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_42384.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Primary sharp EM map | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.828 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Additional map: Primary nonsharpen cryo-EM map
File | emd_42384_additional_1.map | ||||||||||||
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Annotation | Primary nonsharpen cryo-EM map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: Sharpen EM map by DeepEMhancer
File | emd_42384_additional_2.map | ||||||||||||
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Annotation | Sharpen EM map by DeepEMhancer | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: EM half map B
File | emd_42384_half_map_1.map | ||||||||||||
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Annotation | EM half map B | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: EM half map A
File | emd_42384_half_map_2.map | ||||||||||||
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Annotation | EM half map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : the human clamp-clamp loader complex PCNA-CTF18
+Supramolecule #1: the human clamp-clamp loader complex PCNA-CTF18
+Macromolecule #1: Chromosome transmission fidelity protein 18 homolog
+Macromolecule #2: Replication factor C subunit 2
+Macromolecule #3: Replication factor C subunit 5
+Macromolecule #4: Replication factor C subunit 4
+Macromolecule #5: Replication factor C subunit 3
+Macromolecule #6: Proliferating cell nuclear antigen
+Macromolecule #7: MAGNESIUM ION
+Macromolecule #8: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER
+Macromolecule #9: ADENOSINE-5'-DIPHOSPHATE
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.3 mg/mL |
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Buffer | pH: 7.5 |
Grid | Model: Quantifoil R2/1 / Support film - Material: GOLD / Support film - topology: HOLEY |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 280 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.9000000000000001 µm / Nominal defocus min: 1.2 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Overall B value: 123 |
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Output model | PDB-8umu: |