title: A novel nucleoside hydrolase from Lactobacillus buchneri LBK78 catalyzing hydrolysis of 2'-O-methylribonucleosides.
abstract: 2'-O-Methylribonucleosides (2'-OMe-NRs) are promising raw materials for nucleic acid drugs because of their high thermal stability and nuclease tolerance. In the course of microbial screening for metabolic activity toward 2'-OMe-NRs, Lactobacillus buchneri LBK78 was found to decompose 2'-O-methyluridine (2'-OMe-UR). The enzyme responsible was partially purified from L. buchneri LBK78 cells by a four-step purification procedure, and identified as a novel nucleoside hydrolase. This enzyme, LbNH, belongs to the nucleoside hydrolase superfamily, and formed a homotetrameric structure composed of subunits with a molecular mass around 34 kDa. LbNH hydrolyzed 2'-OMe-UR to 2'-O-methylribose and uracil, and the kinetic constants were Km of 0.040 mM, kcat of 0.49 s(-1), and kcat/Km of 12 mM(-1) s(-1). In a substrate specificity analysis, LbNH preferred ribonucleosides and 2'-OMe-NRs as its hydrolytic substrates, but reacted weakly with 2'-deoxyribonucleosides. In a phylogenetic analysis, LbNH showed a close relationship with purine-specific nucleoside hydrolases from trypanosomes.
authors.affiliation: a Division of Applied Life Sciences, Graduate School of Agriculture , Kyoto University , Sakyo-ku, Kyoto , Japan.
authors.forename: Yuuki
authors.initials: Y
authors.lastname: Mitsukawa
authors.affiliation: b Industrial Microbiology, Graduate School of Agriculture , Kyoto University , Sakyo-ku, Kyoto , Japan.
authors.forename: Makoto
authors.initials: M
authors.lastname: Hibi
authors.affiliation: a Division of Applied Life Sciences, Graduate School of Agriculture , Kyoto University , Sakyo-ku, Kyoto , Japan.
authors.forename: Narihiro
authors.initials: N
authors.lastname: Matsutani
authors.affiliation: a Division of Applied Life Sciences, Graduate School of Agriculture , Kyoto University , Sakyo-ku, Kyoto , Japan.
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Horinouchi
authors.affiliation: b Industrial Microbiology, Graduate School of Agriculture , Kyoto University , Sakyo-ku, Kyoto , Japan.
authors.forename: Satomi
authors.initials: S
authors.lastname: Takahashi
authors.affiliation: a Division of Applied Life Sciences, Graduate School of Agriculture , Kyoto University , Sakyo-ku, Kyoto , Japan.
authors.forename: Jun
authors.initials: J
authors.lastname: Ogawa
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D003001:Cloning, Molecular; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D007700:Kinetics; D007778:Lactobacillus / Q000145:classification / Q000201:enzymology* / Q000235:genetics; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D055503:Protein Multimerization; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012266:Ribose / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D014498:Uracil / Q000737:chemistry / Q000378:metabolism; D014529:Uridine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; C015362:D-2-O-methylribose; C052202:2'-O-methyluridine; D014498:Uracil; D012266:Ribose; D009699:N-Glycosyl Hydrolases; D014529:Uridine
keywords: 2′-O-methylribonucleosides; Lactobacilli; nucleic acid drugs; nucleoside hydrolase