8DQZ

Intermediate state of RFC:PCNA bound to a 3' ss/dsDNA junction

Summary for 8DQZ

• Entry DOI: 10.2210/pdb8dqz/pdb
• EMDB information: 27662 27663 27666 27667 27668 27669 27670 27671 27672 27673
• Descriptor: Replication factor C subunit 1, MAGNESIUM ION, GUANOSINE-5'-DIPHOSPHATE, ... (11 entities in total)
• Functional Keywords: replication-dna complex, replication/dna
• Biological source: Saccharomyces cerevisiae (baker's yeast); More
• Total number of polymer chains: 10
• Total formula weight: 363787.82

Authors

Schrecker, M.,Hite, R.K. (deposition date: 2022-07-20, release date: 2022-08-24, Last modification date: 2024-02-14)

Primary citation

Schrecker, M.,Castaneda, J.C.,Devbhandari, S.,Kumar, C.,Remus, D.,Hite, R.K.
Multistep loading of a DNA sliding clamp onto DNA by replication factor C.
Elife, 11:-, 2022

PubMed Abstract: The DNA sliding clamp proliferating cell nuclear antigen (PCNA) is an essential co-factor for many eukaryotic DNA metabolic enzymes. PCNA is loaded around DNA by the ATP-dependent clamp loader replication factor C (RFC), which acts at single-stranded (ss)/double-stranded DNA (dsDNA) junctions harboring a recessed 3' end (3' ss/dsDNA junctions) and at DNA nicks. To illuminate the loading mechanism we have investigated the structure of RFC:PCNA bound to ATPγS and 3' ss/dsDNA junctions or nicked DNA using cryogenic electron microscopy. Unexpectedly, we observe open and closed PCNA conformations in the RFC:PCNA:DNA complex, revealing that PCNA can adopt an open, planar conformation that allows direct insertion of dsDNA, and raising the question of whether PCNA ring closure is mechanistically coupled to ATP hydrolysis. By resolving multiple DNA-bound states of RFC:PCNA we observe that partial melting facilitates lateral insertion into the central channel formed by RFC:PCNA. We also resolve the Rfc1 N-terminal domain and demonstrate that its single BRCT domain participates in coordinating DNA prior to insertion into the central RFC channel, which promotes PCNA loading on the lagging strand of replication forks in vitro. Combined, our data suggest a comprehensive and fundamentally revised model for the RFC-catalyzed loading of PCNA onto DNA.

PubMed: 35939393
DOI: 10.7554/eLife.78253

Experimental method

ELECTRON MICROSCOPY (2.92 Å)