The archaeal Hjm helicase has recQ‐like functions, and may be involved in repair of stalled replication fork

Abstract

The archaeal Hjm is a structure-specific DNA helicase, which was originally identified in the hyperthermophilic archaeon, Pyrococcus furiosus , by in vitro screening for Holliday junction migration activity. Further biochemical analyses of the Hjm protein from P. furiosus showed that this protein preferably binds to fork-related Y-structured DNAs and unwinds their double-stranded regions in vitro , just like the E. coli RecQ protein. Furthermore, genetic analyses showed that Hjm produced in E. coli cells partially complemented the defect of functions of RecQ in a recQ mutant E. coli strain. These results suggest that Hjm may be a functional counterpart of RecQ in Archaea, in which it is necessary for the maintenance of genome integrity, although the amino acid sequences are not conserved. The functional interaction of Hjm with PCNA for its helicase activity further suggests that the Hjm works at stalled replication forks, as a member of the reconstituted replisomes to restart replication.

Hjm/Hel308A DNA Helicase fromSulfolobus tokodaiiPromotes Replication Fork Regression and Interacts with Hjc Endonuclease In Vitro

“…3B and C), perhaps due to the fact that the leading nascent strand of the replication fork resembles a 5Ј overhang structure, while the lagging nascent strand of the replication fork resembles a 3Ј overhang structure. This unwinding activity of StoHjm differs from those reported for Hjm from P. furiosus and for Hel308a from M. thermautotrophicus (9,11), as these enzymes unwind only the lagging strand of the replication fork and the 3Ј overhang structure. The difference in unwinding activity between StoHjm and Hjm/Hel308a may be due to evolutional divergence between Euryarchaeota and Crenarchaeota and may indicate divergent cellular functions of Hjm/Hel308a helicases in these two major archaeal subdomains.…”

“…These results indicate that StoHjm preferentially binds to ssDNA, overhang structures, and branched DNA but not blunt-ended DNA. The DNA binding properties of StoHjm are similar to those of Hjm from P. furiosus and Hel308a from M. thermautotrophicus (9,11). StoHjm preferentially unwinds nascent strands of the replication fork.…”

“…There have been a few reports about DNA helicases that are involved in recombination repair in archaea (8,9,11,13) but none regarding helicases that promote fork regression. Two helicases, Holliday junction migration DNA helicase (Hjm) from Pyrococcus furiosus and Hel308a from Methano-thermobacter thermautotrophicus, have been identified recently (8,9,11). However, the real functions of Hjm/Hel308a helicases are unknown.…”

Genetic analysis of DNA repair in the hyperthermophilic archaeon, Thermococcus kodakaraensis

“…In addition to the Δhef mutant, the sensitivity of Δhjm to MMC is obviously higher than that of the other three deletion mutants, and therefore, the Hjm helicase probably functions in the repair of stalled replication forks, especially those caused by ICLs. The substrate specificity of the P. furiosus Hjm helicase in vitro is consistent with this prediction (Fujikane et al, 2006). It would be interesting to determine how Hef (especially its helicase activity) and Hjm function together in stalled replication fork repair in the cells.…”

“…A biochemical screen to detect an activity that can promote the migration of synthetic Holliday junctions identified the Hjm/Hel308a helicase (Fujikane et al, 2005). This archaeal protein partially complemented the defect of RecQ functions in a recQ mutant Escherichia coli cells, and may be involved in stalled replication fork stabilization like eukaryotic RecQ family proteins including WRN, BLM, and Sgs1 (Fujikane et al, 2006;Guy and Bolt, 2005). In vitro biochemical and structural analyses of these proteins in Archaea have been performed, and their in vitro activities have been characterized.…”