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TitleDNA is loaded through the 9-1-1 DNA checkpoint clamp in the opposite direction of the PCNA clamp.
Journal, issue, pagesNat Struct Mol Biol, Vol. 29, Issue 4, Page 376-385, Year 2022
Publish dateMar 21, 2022
AuthorsFengwei Zheng / Roxana E Georgescu / Nina Y Yao / Michael E O'Donnell / Huilin Li /
PubMed AbstractThe 9-1-1 DNA checkpoint clamp is loaded onto 5'-recessed DNA to activate the DNA damage checkpoint that arrests the cell cycle. The 9-1-1 clamp is a heterotrimeric ring that is loaded in ...The 9-1-1 DNA checkpoint clamp is loaded onto 5'-recessed DNA to activate the DNA damage checkpoint that arrests the cell cycle. The 9-1-1 clamp is a heterotrimeric ring that is loaded in Saccharomyces cerevisiae by Rad24-RFC (hRAD17-RFC), an alternate clamp loader in which Rad24 replaces Rfc1 in the RFC1-5 clamp loader of proliferating cell nuclear antigen (PCNA). The 9-1-1 clamp loading mechanism has been a mystery, because, unlike RFC, which loads PCNA onto a 3'-recessed junction, Rad24-RFC loads the 9-1-1 ring onto a 5'-recessed DNA junction. Here we report two cryo-EM structures of Rad24-RFC-DNA with a closed or 27-Å open 9-1-1 clamp. The structures reveal a completely unexpected mechanism by which a clamp can be loaded onto DNA. Unlike RFC, which encircles DNA, Rad24 binds 5'-DNA on its surface, not inside the loader, and threads the 3' ssDNA overhang into the 9-1-1 clamp from above the ring.
External linksNat Struct Mol Biol / PubMed:35314830 / PubMed Central
MethodsEM (single particle)
Resolution3.17 - 3.23 Å
Structure data

25121

7sgz

EMDB-25121: Cryo-EM 3D map of the yeast Rad24-RFC loader bound to DNA and the closed 9-1-1 clamp
PDB-7sgz: Structure of the yeast Rad24-RFC loader bound to DNA and the closed 9-1-1 clamp
Method: EM (single particle) / Resolution: 3.17 Å

25122

7sh2

EMDB-25122: Cryo-EM 3D map of the yeast Rad24-RFC loader bound to DNA and the open 9-1-1 clamp
PDB-7sh2: Structure of the yeast Rad24-RFC loader bound to DNA and the open 9-1-1 clamp
Method: EM (single particle) / Resolution: 3.23 Å

Chemicals

ChemComp-AGS:
PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / ATP-gamma-S, energy-carrying molecule analogue*YM

ChemComp-MG:
Unknown entry

ChemComp-ADP:
ADENOSINE-5'-DIPHOSPHATE / ADP, energy-carrying molecule*YM / Adenosine diphosphate

ChemComp-PO4:
PHOSPHATE ION / Phosphate

Source
  • synthetic construct (others)
  • saccharomyces cerevisiae (brewer's yeast)
KeywordsDNA BINDING PROTEIN/DNA / DNA damage repair / Rad24-RFC / 9-1-1 clamp / DNA clamp / alternative clamp loader / DNA damage signaling / DNA BINDING PROTEIN-DNA complex

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