Crystal Structure of the HRDC Domain of Human Werner Syndrome Protein, WRN

Ken Kitano,Toshio Hakoshima,Nozomi Yoshihara

doi:10.1074/jbc.m610142200

Ken Kitano, Toshio Hakoshima + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m610142200

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Abstract

Werner syndrome is a human premature aging disorder characterized by chromosomal instability. The disease is caused by the functional loss of WRN, a member of the RecQ-helicase family that plays an important role in DNA metabolic pathways. WRN contains four structurally folded domains comprising an exonuclease, a helicase, a winged-helix, and a helicase-and-ribonuclease D/C-terminal (HRDC) domain. In contrast to the accumulated knowledge pertaining to the biochemical functions of the three N-terminal domains, the function of C-terminal HRDC remains unknown. In this study, the crystal structure of the human WRN HRDC domain has been determined. The domain forms a bundle of alpha-helices similar to those of Saccharomyces cerevisiae Sgs1 and Escherichia coli RecQ. Surprisingly, the extra ten residues at each of the N and C termini of the domain were found to participate in the domain architecture by forming an extended portion of the first helix alpha1, and a novel looping motif that traverses straight along the domain surface, respectively. The motifs combine to increase the domain surface of WRN HRDC, which is larger than that of Sgs1 and E. coli. In WRN HRDC, neither of the proposed DNA-binding surfaces in Sgs1 or E. coli is conserved, and the domain was shown to lack DNA-binding ability in vitro. Moreover, the domain was shown to be thermostable and resistant to protease digestion, implying independent domain evolution in WRN. Coupled with the unique long linker region in WRN, the WRN HRDC may be adapted to play a distinct function in WRN that involves protein-protein interactions.

Highlights

Rosis, arteriosclerosis, type II diabetes, cataracts, and a marked increase in the incidence of cancer [1, 2]
Domain Characterization and Crystallization—To date, the purification and structure determination of RecQ family helicase-and-ribonuclease D/C-terminal (HRDC) domains has only been reported for two unicellular proteins, S. cerevisiae Sgs1 [19] and E. coli RecQ [20], and there have been no reports on the successful isolation and purification of the HRDC domains from mammalian RecQ proteins
In this report, we presented the first structure of the human Werner syndrome protein (WRN) HRDC domain

Summary

Introduction

Rosis, arteriosclerosis, type II diabetes, cataracts, and a marked increase in the incidence of cancer [1, 2]. Detailed studies of core fragments from Escherichia coli RecQ [15, 16] and BLM [17] demonstrated that the fragment is sufficient for ATPase and DNA unwinding activities in vitro, whereas the function of the C-terminal HRDC domain has not been elucidated. Structure of WRN HRDC able of the RecQ family domains (WRN HRDC region shares only ϳ20% homology with Sgs and E. coli RecQ) with examples of human proteins that lack the HRDC domain altogether, including RecQ1, RecQ4, and all isoforms of RecQ5. WRN is distinct from E. coli RecQ in having a considerably long linker region (Ͼ70 residues) that connects HRDC to the N-terminal winged-helix domain. In an effort to delineate the biochemical function of HRDC in WRN, we have determined the crystal structure of the human WRN HRDC domain This represents the first structure of a mammalian RecQ family HRDC domain. The domain was further shown to be stable in solution and lack DNA binding ability

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Discussion

Conclusion