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- PDB-8fs7: Structure of S. cerevisiae Rad24-RFC loading the 9-1-1 clamp onto... -
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Open data
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Basic information
Entry | Database: PDB / ID: 8fs7 | ||||||||||||
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Title | Structure of S. cerevisiae Rad24-RFC loading the 9-1-1 clamp onto a 10-nt gapped DNA in step 5 (closed 9-1-1 and stably bound chamber DNA) | ||||||||||||
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![]() | CELL CYCLE/DNA / DNA damage repair / Rad24-RFC / 9-1-1 clamp / DNA clamp / alternative clamp loader / DNA damage signaling / DNA BINDING PROTEIN-DNA complex / CELL CYCLE-DNA complex | ||||||||||||
Function / homology | ![]() meiotic DNA integrity checkpoint signaling / checkpoint clamp complex / DNA clamp unloading / Gap-filling DNA repair synthesis and ligation in GG-NER / Rad17 RFC-like complex / Ctf18 RFC-like complex / DNA clamp loader activity / DNA replication factor C complex / Elg1 RFC-like complex / Polymerase switching ...meiotic DNA integrity checkpoint signaling / checkpoint clamp complex / DNA clamp unloading / Gap-filling DNA repair synthesis and ligation in GG-NER / Rad17 RFC-like complex / Ctf18 RFC-like complex / DNA clamp loader activity / DNA replication factor C complex / Elg1 RFC-like complex / Polymerase switching / Translesion Synthesis by POLH / telomere maintenance via recombination / Translesion synthesis by REV1 / Translesion synthesis by POLK / Translesion synthesis by POLI / DNA replication checkpoint signaling / Activation of ATR in response to replication stress / Termination of translesion DNA synthesis / mitotic DNA replication checkpoint signaling / reciprocal meiotic recombination / sister chromatid cohesion / recombinational repair / mitotic intra-S DNA damage checkpoint signaling / mitotic sister chromatid cohesion / leading strand elongation / Gap-filling DNA repair synthesis and ligation in TC-NER / Dual incision in TC-NER / subtelomeric heterochromatin formation / meiotic cell cycle / mismatch repair / 3'-5' exonuclease activity / telomere maintenance / DNA damage checkpoint signaling / cellular response to ionizing radiation / nucleotide-excision repair / double-strand break repair via homologous recombination / DNA-templated DNA replication / double-strand break repair / double-stranded DNA binding / site of double-strand break / chromosome, telomeric region / damaged DNA binding / DNA repair / chromatin binding / ATP hydrolysis activity / DNA binding / ATP binding / nucleus / cytosol Similarity search - Function | ||||||||||||
Biological species | ![]() ![]() synthetic construct (others) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.85 Å | ||||||||||||
![]() | Zheng, F. / Georgescu, R. / Yao, Y.N. / O'Donnell, M.E. / Li, H. | ||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Structures of 9-1-1 DNA checkpoint clamp loading at gaps from start to finish and ramification to biology. Authors: Fengwei Zheng / Roxana E Georgescu / Nina Y Yao / Michael E O'Donnell / Huilin Li / ![]() Abstract: Recent structural studies show the Rad24-RFC loads the 9-1-1 checkpoint clamp onto a recessed 5' end by binding the 5' DNA on Rad24 at an external surface site and threading the 3' ssDNA into the ...Recent structural studies show the Rad24-RFC loads the 9-1-1 checkpoint clamp onto a recessed 5' end by binding the 5' DNA on Rad24 at an external surface site and threading the 3' ssDNA into the well-established internal chamber and into 9-1-1. We find here that Rad24-RFC loads 9-1-1 onto DNA gaps in preference to a recessed 5' DNA end, thus presumably leaving 9-1-1 on a 3' ss/ds DNA after Rad24-RFC ejects from the 5' gap end and may explain reports of 9-1-1 directly functioning in DNA repair with various TLS polymerases, in addition to signaling the ATR kinase. To gain a deeper understanding of 9-1-1 loading at gaps we report high-resolution structures of Rad24-RFC during loading of 9-1-1 onto 10-nt and 5-nt gapped DNAs. At a 10-nt gap we captured five Rad24-RFC-9-1-1 loading intermediates in which the 9-1-1 DNA entry gate varies from fully open to fully closed around DNA using ATPγS, supporting the emerging view that ATP hydrolysis is not needed for clamp opening/closing, but instead for dissociation of the loader from the clamp encircling DNA. The structure of Rad24-RFC-9-1-1 at a 5-nt gap shows a 180° axially rotated 3'-dsDNA which orients the template strand to bridge the 3'- and 5'- junctions with a minimum 5-nt ssDNA. The structures reveal a unique loop on Rad24 that limits the length of dsDNA in the inner chamber, and inability to melt DNA ends unlike RFC, thereby explaining Rad24-RFC's preference for a preexisting ssDNA gap and suggesting a direct role in gap repair in addition to its checkpoint role. | ||||||||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 575.4 KB | Display | ![]() |
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PDB format | ![]() | 450.2 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.9 MB | Display | ![]() |
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Full document | ![]() | 1.9 MB | Display | |
Data in XML | ![]() | 86.9 KB | Display | |
Data in CIF | ![]() | 128.8 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 29416MC ![]() 8fs3C ![]() 8fs4C ![]() 8fs5C ![]() 8fs6C ![]() 8fs8C M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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1 |
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Components
-Protein , 2 types, 2 molecules AH
#1: Protein | Mass: 62792.496 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: RAD24, YER173W, SYGP-ORF60 / Production host: ![]() ![]() |
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#8: Protein | Mass: 69787.500 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: DDC1 / Production host: ![]() ![]() |
-Replication factor C subunit ... , 4 types, 4 molecules BCDE
#2: Protein | Mass: 36201.039 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: RFC4, YOL094C, O0923 / Production host: ![]() ![]() |
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#3: Protein | Mass: 37841.051 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: RFC3, YNL290W, N0533 / Production host: ![]() ![]() |
#4: Protein | Mass: 39794.473 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: RFC2, YJR068W, J1808 / Production host: ![]() ![]() |
#5: Protein | Mass: 39993.582 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: RFC5, YBR087W, YBR0810 / Production host: ![]() ![]() |
-DNA damage checkpoint control protein ... , 2 types, 2 molecules FG
#6: Protein | Mass: 53207.797 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: MEC3, PIP3, PSO9, YLR288C, L8003.15 / Production host: ![]() ![]() |
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#7: Protein | Mass: 45637.527 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: RAD17, GI527_G0005737 / Production host: ![]() ![]() |
-DNA chain , 1 types, 1 molecules I
#9: DNA chain | Mass: 15389.855 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
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-Primer strand ... , 2 types, 2 molecules JK
#10: DNA chain | Mass: 6464.172 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
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#11: DNA chain | Mass: 6084.940 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
-Non-polymers , 3 types, 9 molecules ![](data/chem/img/MG.gif)
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#12: Chemical | ChemComp-MG / #13: Chemical | ChemComp-AGS / #14: Chemical | ChemComp-ADP / | |
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-Details
Has ligand of interest | N |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
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Source (natural) |
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Source (recombinant) | Organism: ![]() ![]() | ||||||||||||||||||||||||||||
Buffer solution | pH: 7.5 | ||||||||||||||||||||||||||||
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||
Vitrification | Cryogen name: ETHANE |
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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: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 1900 nm / Nominal defocus min: 1300 nm |
Image recording | Electron dose: 64 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
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Processing
Software | Name: PHENIX / Version: 1.20.1_4487: / Classification: refinement | ||||||||||||||||||||||||
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CTF correction | Type: NONE | ||||||||||||||||||||||||
3D reconstruction | Resolution: 2.85 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 292725 / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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