Examination of the roles of a conserved motif in the PriA helicase in structure-specific DNA unwinding and processivity.

Alexander T Duckworth,James L Keck,Tricia A Windgassen,Andrei Chernov

doi:10.1371/journal.pone.0255409

Alexander T Duckworth, James L Keck + Show 2 more

Open Access

https://doi.org/10.1371/journal.pone.0255409

Copy DOI

Abstract

DNA replication complexes (replisomes) frequently encounter barriers that can eject them prematurely from the genome. To avoid the lethality of incomplete DNA replication that arises from these events, bacteria have evolved “DNA replication restart” mechanisms to reload replisomes onto abandoned replication forks. The Escherichia coli PriA DNA helicase orchestrates this process by recognizing and remodeling replication forks and recruiting additional proteins that help to drive replisome reloading. We have identified a conserved sequence motif within a linker region of PriA that docks into a groove on the exterior of the PriA helicase domain. Alterations to the motif reduce the apparent processivity and attenuate structure-specific helicase activity in PriA, implicating the motif as a potential autoregulatory element in replication fork processing. The study also suggests that multiple PriA molecules may function in tandem to enhance DNA unwinding processivity, highlighting an unexpected similarity between PriA and other DNA helicases.

Highlights

The essential process of genome duplication is catalyzed by protein complexes called replisomes
Examination of the crystal structures of Klebsiella pneumoniae and E. coli PriA [6, 9] showed that Trp186 and two less well conserved residues (Arg187 and Tyr190 in E. coli PriA) form a motif that docks into a conserved pocket on the PriA helicase domain (HD) (Fig 1C)
The position of the binding pocket in the HD and the proximity of the linker motif to the 30BD and WH domains led us to hypothesize that the linker/HD interaction may be important for regulating PriA activity

Summary

Introduction

The essential process of genome duplication is catalyzed by protein complexes called replisomes. Replisome dissociation halts replication and leaves behind abandoned replication forks [1,2,3]. To rescue prematurely arrested DNA replication processes in these instances, cells have evolved DNA replication restart mechanisms that reload replisomes onto abandoned replication forks. The PriA DNA helicase is a central component of the primosome and functions to recognize and remodel abandoned replication forks for replisome reloading [6,7,8]. PriA initiates replication restart by binding to abandoned replication forks in a structure-dependent manner. This allows PriA to act in a sequence-independent fashion, which is important because replication fork abandonment can happen anywhere along a chromosome. PriA structure specificity orients the helicase to unwind lagging strand DNA.

Methods

Results

Conclusion