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Data from: Domain dynamics within the PriA DNA helicase regulate DNA replication restart

Deorio, Haley1; Duckworth, Alexander2; Forsythe, Gavin1; Sung, Andrew3; Grant, Tim1; Sandler, Steven4; Keck, James1

Research facility: University of Wisconsin–Madison
Published Sep 07, 2026 on Dryad. https://doi.org/10.5061/dryad.wpzgmsc40

Data files

Sep 07, 2026 version files 1.59 GB

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Abstract

Prematurely terminated DNA replication processes in bacteria must be restarted for successful genome duplication. In Escherichia coli, the PriA DNA helicase orchestrates replication restart by assembling the PriA/PriB/DnaT preprimosome complex at abandoned DNA replication forks and reloading the replicative machinery. We show that the structure of the lagging strand—whether single- or double-stranded—influences the position of the cysteine-rich region in PriA (PriACRR), thereby regulating PriA biochemical activity and preprimosome formation. When PriA binds to replication forks with a single-stranded lagging strand, the PriACRR is found either in a state similar to free PriA or in a rotated position that encircles the lagging strand and allows PriB recruitment. Binding to a replication fork with a duplex lagging strand neither the PriACRR position nor promotes PriB binding, suggesting PriA must unwind duplex lagging-strand DNA to trigger PriACRR movement and subsequent preprimosome formation. PriA variants designed to destabilize the two PriACRR positions differentially affect ATPase activity, helicase function, and PriB binding in vitro, and PriA-mediated replication restart in vivo. These results demonstrate that the PriACRR functions as a regulatory switch that detects the structure of the lagging strand and controls the biochemical and cellular activities of PriA to restart DNA replication.