Biochemical characterization of the WRN-1 RecQ helicase of Caenorhabditis elegans.
The highly conserved RecQ helicases are essential for the maintenance of genomic stability. Werner syndrome protein, WRN, is one of five human RecQ helicase homologues, and a deficiency of the protein causes a hereditary premature aging disorder that is characterized by genomic instability. A WRN orthologue, wrn-1 lacking the exonuclease domain, has been identified in the nematode Caenorhabditis elegans. wrn-1(RNAi) in C. elegans has a shortened life span, increased sensitivity to DNA damage, and accelerated aging phenotypes. However, little is known about its enzymatic activity. We purified the recombinant C. elegans WRN-1 protein (CeWRN-1) and then investigated its substrate specificity in vitro to improve our understanding of its function in vivo. We found that CeWRN-1 is an ATP-dependent 3'-5' helicase capable of unwinding a variety of DNA structures such as forked duplexes, Holliday junctions, bubble substrates, D-loops, and flap duplexes, and 3'-tailed duplex substrates. Distinctly, CeWRN-1 is able to unwind a long forked duplex compared to human WRN. Furthermore, CeWRN-1 helicase activity on a long DNA duplex is stimulated by C. elegans replication protein A (CeRPA) that is shown to interact with CeWRN-1 by a dot blot. The ability of CeWRN-1 to unwind these DNA structures may improve the access for DNA repair and replication proteins that are important for preventing the accumulation of abnormal structures, contributing to genomic stability.
Hyun
Moonjung
M
Department of Life Sciences, University of Ulsan, Ulsan 680749, Korea.
Bohr
Vilhelm A
VA
Ahn
Byungchan
B
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D017173:Caenorhabditis elegans / Q000201:enzymology*; D029742:Caenorhabditis elegans Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017124:Conserved Sequence; D004247:DNA / Q000235:genetics; D004249:DNA Damage; D004265:DNA Helicases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004260:DNA Repair; D004261:DNA Replication; D005838:Genotype; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity
D029742:Caenorhabditis elegans Proteins; D011994:Recombinant Proteins; D004247:DNA; D000251:Adenosine Triphosphatases; D004265:DNA Helicases; C488009:WRN-1 protein, C elegans