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We report the characterization of a uracil-DNA glycosylase(UDG) from the hyperthermophilic archaea Pyrococcus furiosus(P.furiosus).P.furiosus UDG(PfUDG) has high sequence similarity to the families IV and V UDGs(thermostable UDG family and PaUDG-b family).PfUDG excises uracil from various DNA substrates with the following order:U/T≈U/C>U/G≈U/AP≈U/->U/U≈U/I≈U/A.The optimal temperature and pH value for uracil excision by PfUDG are 70 °C and 9.0,respectively.The removal of U is inhibited by the divalent ions of Fe,Ca,Zn,Cu,Co,Ni and Mn,as well as a high concentration of NaCl.The phosphorothioates near uracil strongly inhibit the excision of uracil by PfUDG.Interestingly,pfuDNA(Pyrococcus furiosus DNA) polymerase,which tightly binds the uracil-carrying oligonucleotide,does not inhibit the excision by PfUDG,suggesting PfUDG in vivo functions as the repair enzyme to excise uracil damage in genome.
We report the characterization of a uracil-DNA glycosylase (UDG) from the hyperthermophilic archaea Pyrococcus furiosus (P. furiosus) .P. furiosus UDG (PfUDG) has a high sequence similarity to the families IV and V UDGs (thermostable UDG family and PaUDG- b family). PfUDG excises uracil from various DNA substrates with the following order: U / T≈U / C> U / G≈U / AP≈U / -> U / U≈U / I≈U / A.The optimal temperature and pH value for uracil excision by PfUDG are 70 ° C and 9.0, respectively. The removal of U is inhibited by the divalent ions of Fe, Ca, Zn, Cu, Co, Ni and Mn, as well as a high concentration of NaCl. The phosphorothioates near uracil strongly inhibit the excision of uracil by PfUDG.Interestingly, pfuDNA (Pyrococcus furiosus DNA) polymerase, which tightly binds the uracil-carrying oligonucleotide, does not inhibit the excision by PfUDG, suggesting PfUDG in vivo functions as the repair enzyme to excise uracil damage in genome.