title: Biochemical characterization of endonuclease V from the hyperthermophilic archaeon, Pyrococcus furiosus.
abstract: Endonuclease V (Endo V) is a DNA repair enzyme that recognizes deoxyinosine and cleaves the second phosphodiester bond on the 3' side of the deaminated base lesion. A database search revealed the presence of homologous genes for Endo V in most archaeal species, but the absence in some methanogenic species. We cloned a gene encoding the sequence homologous to Escherichia coli Endo V from the genome of the hyperthermophilic euryarchaeon, Pyrococcus furiosus and purified gene product (PfuEndoV) to homogeneity. In vitro characterization showed that PfuEndoV possesses specific endonuclease activity for the deoxyinosine-containing DNA strand. The activity of the enzyme was maximal at 90°C. Stable complex formation between PfuEndoV and nicked DNA produced by the cleavage reaction was detected by gel mobility shift assays. The molecular mechanisms of the inosine repair pathway including Endo V in the archaeal cells are discussed. Interestingly, PfuEndoV cleaved inosine-containing RNA strands as well as DNA substrates. PfuEndoV may also be involved in RNA metabolism.
authors.affiliation: Department of Bioscience and Biotechnology, Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka, Fukuoka 812-8581, Japan.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Kiyonari
authors.forename: Yuriko
authors.initials: Y
authors.lastname: Egashira
authors.forename: Sonoko
authors.initials: S
authors.lastname: Ishino
authors.forename: Yoshizumi
authors.initials: Y
authors.lastname: Ishino
mesh_terms: D000595:Amino Acid Sequence; D043224:Deoxyribonuclease (Pyrimidine Dimer) / Q000235:genetics / Q000378:metabolism*; D004795:Enzyme Stability; D007288:Inosine / Q000031:analogs & derivatives; D008274:Magnesium / Q000378:metabolism; D008969:Molecular Sequence Data; D020027:Pyrococcus furiosus / Q000201:enzymology* / Q000235:genetics; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D011994:Recombinant Proteins; D007288:Inosine; D012313:RNA; D043224:Deoxyribonuclease (Pyrimidine Dimer); C012271:deoxyinosine; D008274:Magnesium
keywords: DNA repair; Endo V; Pyrococcus furiosus; archaea; hypoxanthine