Yeast DNA-repair gene RAD14 encodes a zinc metalloprotein with affinity for ultraviolet-damaged DNA.
Xeroderma pigmentosum (XP) patients suffer from a high incidence of skin cancers due to a defect in excision repair of UV light-damaged DNA. Of the seven XP complementation groups, A-G, group A represents a severe and frequent form of the disease. The Saccharomyces cerevisiae RAD14 gene is a homolog of the XP-A correcting (XPAC) gene. Like XP-A cells, rad14-null mutants are defective in the incision step of excision repair of UV-damaged DNA. We have purified RAD14 protein to homogeneity from extract of a yeast strain genetically tailored to overexpress RAD14. As determined by atomic emission spectroscopy, RAD14 contains one zinc atom. We also show in vitro that RAD14 binds zinc but does not bind other divalent metal ions. In DNA mobility-shift assays, RAD14 binds specifically to UV-damaged DNA. Removal of cyclobutane pyrimidine dimers from damaged DNA by enzymatic photoreactivation has no effect on binding, strongly suggesting that RAD14 recognizes pyrimidine(6-4)pyrimidone photoproduct sites. These findings indicate that RAD14 functions in damage recognition during excision repair.
Guzder
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Department of Biology, University of Rochester, NY 14627.
Sung
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Prakash
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Prakash
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D000595:Amino Acid Sequence; D004249:DNA Damage; D004260:DNA Repair / Q000235:genetics*; D045643:DNA Repair Enzymes; D004255:Deoxyribodipyrimidine Photo-Lyase / Q000378:metabolism; D004307:Dose-Response Relationship, Radiation; D004926:Escherichia coli / Q000201:enzymology; D005656:Fungal Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D005800:Genes, Fungal; D005822:Genetic Vectors; D007700:Kinetics; D008667:Metalloproteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D011740:Pyrimidine Dimers / Q000378:metabolism; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism* / Q000528:radiation effects; D029701:Saccharomyces cerevisiae Proteins; D014466:Ultraviolet Rays; D015032:Zinc / Q000378:metabolism*
D005656:Fungal Proteins; D008667:Metalloproteins; D011740:Pyrimidine Dimers; C072975:RAD14 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D004255:Deoxyribodipyrimidine Photo-Lyase; D045643:DNA Repair Enzymes; D015032:Zinc