Mechanistic insights into Lhr helicase function in DNA repair.

Ryan J Buckley,Dina Grohmann,Christopher D O Cooper,Kevin Kramm,Edward L Bolt

doi:10.1042/bcj20200379

Ryan J Buckley, Dina Grohmann + Show 3 more

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https://doi.org/10.1042/bcj20200379

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Abstract

The DNA helicase Large helicase-related (Lhr) is present throughout archaea, including in the Asgard and Nanoarchaea, and has homologues in bacteria and eukaryotes. It is thought to function in DNA repair but in a context that is not known. Our data show that archaeal Lhr preferentially targets DNA replication fork structures. In a genetic assay, expression of archaeal Lhr gave a phenotype identical to the replication-coupled DNA repair enzymes Hel308 and RecQ. Purified archaeal Lhr preferentially unwound model forked DNA substrates compared with DNA duplexes, flaps and Holliday junctions, and unwound them with directionality. Single-molecule FRET measurements showed that binding of Lhr to a DNA fork causes ATP-independent distortion and base-pair melting at, or close to, the fork branchpoint. ATP-dependent directional translocation of Lhr resulted in fork DNA unwinding through the ‘parental’ DNA strands. Interaction of Lhr with replication forks in vivo and in vitro suggests that it contributes to DNA repair at stalled or broken DNA replication.

Highlights

Lhr (Large helicase-related) protein is an ATP-dependent DNA translocase and helicase that forms a distinct group within Superfamily 2 helicases [1,2]
Lhr protein is highly conserved throughout archaea and has sequence homology with DDX damage repair proteins found in humans and other eukaryotes [7]
Lhr proteins form two sub-groups, Lhr and Lhr-Core, the latter including the archaeal proteins of 800–900 amino acids arranged into RecA-like and accessory domains required for helicase activity

Summary

Introduction

Lhr (Large helicase-related) protein is an ATP-dependent DNA translocase and helicase that forms a distinct group within Superfamily 2 helicases [1,2]. Lhr was discovered and named in bacteria [2], in which it is present in eight of ∼30 phyla [2,3]. It is widespread in archaea [4], and the archaeal Lhr is a sequence homologue of the DDX-family of uncharacterized putative helicases found in eukaryotes including in humans [5,6,7]. Archaeal and bacterial Lhr proteins show high amino acid sequence identity (typically ∼30%) between their N-terminal 800–900 amino acids, which is referred to as the ‘Lhr-Core’, that comprises their helicase domains [8]. A crystal structure of bacterial Lhr-Core highlights significant similarities with the archaeal DNA repair helicase Hel308 [9,11], most notably in the orientation and interaction of its winged helix domain (WHD) with RecA-like domains typical of Ski2-like helicases [12,13]

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