title: A structure-specific endonuclease from cauliflower (Brassica oleracea var. botrytis) inflorescence.
abstract: A protein with structure-specific endonuclease activity has been purified to near homogeneity from cauliflower ( Brassica oleracea var. botrytis) inflorescence through five successive column chromatographies. The protein is a single polypeptide with a molecular mass of 40 kDa. Using three different branched DNA structures (flap, pseudo-Y and stem-loop) we found that the enzyme, a cauliflower structure-specific endonuclease, cleaved the single-stranded tail in the 5'-flap and 5'-pseudo-Y structures, whereas it could not incise the 3'-flap and 3'-pseudo-Y structures. The incision points occur around the single strand-duplex junction in these DNA substrates and the enzyme leaves 5'-PO4 and 3'-OH termini on DNA. The protein also endonucleolytically cleaves on the 3'-side of the single-stranded region at the junction of unpaired and duplex DNA in the stem-loop structure. The structure-specific endonuclease activity is stimulated by Mg2+ and by Mn2+, but not by Ca2+. Like mammalian FEN-1, the protein has weak 5'-->3' double-stranded DNA-specific exonuclease activity. These results indicate that the cauliflower protein is a plant structure-specific endonuclease like mammalian FEN-1 or may be the plant alternative.
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Science University of Tokyo, 2641 Yamazaki, Noda-shi, Chiba-ken 278, Japan.
authors.forename: S
authors.initials: S
authors.lastname: Kimura
authors.forename: M
authors.initials: M
authors.lastname: Kai
authors.forename: H
authors.initials: H
authors.lastname: Kobayashi
authors.forename: A
authors.initials: A
authors.lastname: Suzuki
authors.forename: H
authors.initials: H
authors.lastname: Morioka
authors.forename: E
authors.initials: E
authors.lastname: Otsuka
authors.forename: K
authors.initials: K
authors.lastname: Sakaguchi
mesh_terms: D001937:Brassica / Q000201:enzymology*; D018744:DNA, Plant / Q000378:metabolism* / Q000648:ultrastructure; D004706:Endodeoxyribonucleases / Q000302:isolation & purification* / Q000378:metabolism; D010940:Plant Proteins / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D018744:DNA, Plant; D010940:Plant Proteins; D004706:Endodeoxyribonucleases