Characterization in vitro and in vivo of the DNA helicase encoded by Deinococcus radiodurans locus DR1572.
Deinococcus radiodurans survives extremely high doses of ionizing and ultraviolet radiation and treatment with various DNA-damaging chemicals. As an effort to identify and characterize proteins that function in DNA repair in this organism, we have studied the protein encoded by locus DR1572. This gene is predicted to encode a Superfamily I DNA helicase, except that genome sequencing indicated that it has a one-base frameshift and would not encode a complete helicase. We have cloned the gene from two different D. radiodurans strains and find that the frameshift mutation is not present. The corrected gene encodes a 755 residue protein that is similar to the Bacillus subtilis YvgS protein and to helicase IV of Escherichia coli. The purified protein (helicase IV(Dr)) has ATP hydrolysis and DNA helicase activity. A truncated protein that lacks 214 residues from the N-terminus, which precede the conserved helicase domain, has greater ATPase activity than the full-length protein but has no detectable helicase activity. Disruption of locus DR1572 in the D. radiodurans chromosome causes greater sensitivity to hydrogen peroxide and methyl-methanesulfonate compared to wild-type cells, but no change in resistance to gamma and ultraviolet radiation and to mitomycin C. The results indicate that locus DR1572 encodes a complete protein that contributes to DNA metabolism in D. radiodurans.
Cao
Zheng
Z
Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, United States.
Julin
Douglas A
DA
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D015139:Blotting, Southern; D003001:Cloning, Molecular; D004249:DNA Damage / Q000187:drug effects* / Q000528:radiation effects*; D004265:DNA Helicases / Q000235:genetics* / Q000378:metabolism*; D004260:DNA Repair / Q000187:drug effects / Q000528:radiation effects; D034301:Deinococcus / Q000201:enzymology* / Q000235:genetics; D005720:Gamma Rays; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006868:Hydrolysis / Q000187:drug effects / Q000528:radiation effects; D066298:In Vitro Techniques; D008741:Methyl Methanesulfonate / Q000494:pharmacology; D016685:Mitomycin / Q000494:pharmacology; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D016877:Oxidants / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D014466:Ultraviolet Rays
D018906:Antineoplastic Agents, Alkylating; D001426:Bacterial Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D016877:Oxidants; D016685:Mitomycin; D000255:Adenosine Triphosphate; D008741:Methyl Methanesulfonate; D006861:Hydrogen Peroxide; D004265:DNA Helicases