Differences in clamp loader mechanism between bacteria and eukaryotes [preprint]
Landeck, Jacob T ; Pajak, Joshua ; Norman, Emily K ; Sedivy, Emma L ; Kelch, Brian A
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Emily K Norman
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Abstract
Clamp loaders are pentameric ATPases that place circular sliding clamps onto DNA, where they function in DNA replication and genome integrity. The central activity of a clamp loader is the opening of the ring-shaped sliding clamp, and the subsequent binding to primer-template (p/t)-junctions. The general architecture of clamp loaders is conserved across all life, suggesting that their mechanism is retained. Recent structural studies of the eukaryotic clamp loader Replication Factor C (RFC) revealed that it functions using a crab-claw mechanism, where clamp opening is coupled to a massive conformational change in the loader. Here we investigate the clamp loading mechanism of the E. coli clamp loader at high resolution using cryo-electron microscopy (cryo-EM). We find that the E. coli clamp loader opens the clamp using a crab-claw motion at a single pivot point, whereas the eukaryotic RFC loader uses motions distributed across the complex. Furthermore, we find clamp opening occurs in multiple steps, starting with a partly open state with a spiral conformation, and proceeding to a wide open clamp in a surprising planar geometry. Finally, our structures in the presence of p/t-junctions illustrate how clamp closes around p/t-junctions and how the clamp loader initiates release from the loaded clamp. Our results reveal mechanistic distinctions in a macromolecular machine that is conserved across all domains of life.
Source
Landeck JT, Pajak J, Norman EK, Sedivy EL, Kelch BA. Differences in clamp loader mechanism between bacteria and eukaryotes. bioRxiv [Preprint]. 2023 Nov 30:2023.11.30.569468. doi: 10.1101/2023.11.30.569468. PMID: 38076975; PMCID: PMC10705477.
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This article is a preprint. Preprints are preliminary reports of work that have not been certified by peer review.