1998 年度実績報告書

RNA触媒機能の分子基盤

研究課題

研究課題/領域番号	09278103
研究機関	京都大学
研究代表者	井上丹京都大学, 大学院・理学研究科, 教授 (40114855)
研究分担者	新田至東京大学, 工学系研究科, 助手 (30272404) 白石英秋京都大学, 大学院・理学研究科, 講師 (90202118) 谷時雄九州大学, 理学部, 助教授 (80197516) 多比良和誠筑波大学, 応用生物化学系, 教授 (10261778) 小松康雄北海道大学, 薬学部, 助手 (30271670)
キーワード	RNA / リボザイム / GNRA-receptor 4 / グループIイントロン / マキシザイム / U6snRNA / RNase P / 23SリボソームRNA
研究概要	・リボザイムの反応の分子機構の解析を、グループIイントロンRNAを材料として行い、そのactivator domainの作用機構及びこれまで未同定のGNRA-receptor motifの存在を明らかにした。また、同RNAの活性中心が存在するdomainの特定に成功した。(井上) ・昨年度報告したヘアピン型リボザイムを、さらに切断活性の高いthree-way型ribozymeを切り出すように変換し、これがヒト誘導型一酸化窒素合成酵素由来のmRNAを部位特異的に切断できることを示した。(小松) ・ 2量体として作用する世界最小のリボザイムを構築することに成功し、さらに、発現システムに工夫を加えることで細胞内において、野生型のリボザイムより高活性を示すマキシザイムの作成に成功した。(多比良) ・ U6snRNAがスプライシング反応の触媒因子として機能する可能性をin vitro反応系で検討したが、有為な結果は得られず、in vivoにおいてさらに機能解析を行うため、U6snRNAに変異を導入した分裂酵母株を作製中である。(谷) ・大腸菌と枯草菌のRNase Pは、活性中心をもつM1 RNAとタンパク質から成る複合体酵素である。それぞれのタンパク質の大量発現系を構築し同酵素の機能構造相関の解析を進めている。また、volvoxの初期発生に関わるG167RNAの解析を開始した。(白石) ・試験管内転写により調製された大腸菌由来の23SリボソームRNAを用いるペプチド結合生成反応の解析をさらに進めている。(新田) ・グアノシンに対するRNAアプタマーの解析を行った.このグアノシン結合RNAの基質特異性の交換には、グアノシン結合RNAのインターナル・ループ内の特定の1個のヌクレオチド残基が関わっている事を明らかにした。(坂本)

研究成果

(24件)

すべてその他

すべて文献書誌 (24件)

[文献書誌] Ikawa,Y.: "Trans-activation of the Tetrahymena ribozyme by its P2-2.1 domains" J.Biochem. 123. 528-533 (1998)
[文献書誌] Naito,Y.: "P5abc of the Tetrahymena ribozyme consists of three functionally independent elements" RNA. 4. 837-846 (1998)
[文献書誌] Ikawa,Y.: "Trans-activation of the Tetrahymena group I intron ribozyme via a non-native RNA-RNA interaction" Nucleic Acids Research. 27・7 (in press). (1999)
[文献書誌] Ikawa,Y.: "A conserved motif in group IC3 introns is a new class of GNRA receptor" Nucleic Acids Research. 27・8 (in press). (1999)
[文献書誌] Komatsu,Y.: "Modification and application of self-trimming hairpin ribozymes to targeting a transcribed RNA in vitro." J.Biochem. 124・5. 986-991 (1998)
[文献書誌] Koizumi,M.: "Design and anti-HIV-1 activity of hammerhead and hairpin ribozymes containing a stable loop." Nucleosides & Nucleotides. (in press).
[文献書誌] Kobayashi,H: "Effets of a high-affinity antibody fragment on DNA polymerase reactions near a (6-4) photoproduct site." Photochem.& Photobio.69・2. 226-230 (1999)
[文献書誌] Ota,N.: "Determination of interactions between structured nucleic acids by fluorescence resonance energy transfer (FRET): selection of target sites for functional nucleic acids" Nucleic Acids Res.26. 735-743 (1998)
[文献書誌] Kawasaki,H.: "p300 and ATF-2 are components of the DRF complex, which regulates retinoicacid-and E1A-mediated transcription of the c-jun gene in F9 cells." Genes & Dev.12. 233-245 (1998)
[文献書誌] Kawasaki,H.: "Distinct roles of the co-activators p300 and CBP in retinoic-acid-induced F9-cell differentiation." Nature. 393. 284-289 (1998)
[文献書誌] Ota,N.: "Effects of helical structures formed by the binding arms of DNAzymes and their substrates on catalytic activity" Nucleic Acids Res.26. 3385-3391 (1998)
[文献書誌] Kuwabara,T.: "Formation of a catalytically active dimer by tRNA Val-driven short ribozymes" Nature Biotechnology. 16. 961-965 (1998)
[文献書誌] Kuwabara,T.: "A novel nallosterically trans-activated ribozyme, the maxizyme, with exceptional specificity in vitro and in vivo." Mol.Cell. 2. 617-627 (1998)
[文献書誌] Kuwabara,T.: "tRNA Val-heterodimeric maxizymes with high potential as gene-inactivating agent: simultaneous cleavage at two sites in HIV-1tat mRNA in cultured cells." Proc.Natl.Acad.Sci.USA96. 1886-1891 (1999)
[文献書誌] Shibuya,T.: "Characterization of the ptr6+ gene in fission yeast: A possible involvement of a transcriptional coactivator TAF in nucleocytoplasmic transport of mRNA" Genetics. in press (1999)
[文献書誌] Kanai,N.: "A “G" to “A" transition at the last nucleotide of exon 6 of the gene(868G-A) may result in either a splice or missense mutation in patients with X-linked severe combined immunodeficiency" Human Genet.104. 36-42 (1999)
[文献書誌] Habara,Y.: "The fission yeast prp10+ gene involved in pre-mRNA splicing encodes a homologue of highly conserved splicing factor, SAP155" Nucleic Acids Res.26・24. 5662-5669 (1998)
[文献書誌] Imamura,O: "Cloning and characterization of a human DEAH-box RNA helicase, a functional homolog of fission yeast Cdc28/Prp8." Nucleic Acids Res.26. 2063-2068 (1998)
[文献書誌] Nitta,I.: "Possible involvement of Escherichia coli 23S ribosomal RNA in peptide bond formation." RNA. 4. 257-267 (1998)
[文献書誌] Nitta,I.: "Reconstitution of peptide bond formation with Escherichia coli 23S ribosomal RNA domains." Science. 281. 666-669 (1998)
[文献書誌] Nitta,I.: "A novel cell-free system for peptide synthesis driven by pyridine." Biol.Chem.379. 819-829 (1998)
[文献書誌] Kimoto,M.: "RNA aptamers that specifically bind to the Ras-binding domain of Raf-1" FEBS Letters. 441. 322-326 (1998)
[文献書誌] Kiga,D.: "An RNA aptamer to the xanthine/guanine base with a distictive mode of purine recognition" Nucleic Acids Research. 26・7. 1755-1760 (1998)
[文献書誌] Seio,K.: "Chemical synthesis and properties of conformationally fixed diuridine monophosphates as builiding blocks of the RNA turn motif" Journal of Organic Chemistry. 63・5. 1429-1443 (1998)

1998 年度 実績報告書

RNA触媒機能の分子基盤

研究代表者

井上 丹 京都大学, 大学院・理学研究科, 教授 (40114855)

研究成果

[文献書誌] Ikawa,Y.: "Trans-activation of the Tetrahymena ribozyme by its P2-2.1 domains" J.Biochem. 123. 528-533 (1998)

[文献書誌] Naito,Y.: "P5abc of the Tetrahymena ribozyme consists of three functionally independent elements" RNA. 4. 837-846 (1998)

[文献書誌] Ikawa,Y.: "Trans-activation of the Tetrahymena group I intron ribozyme via a non-native RNA-RNA interaction" Nucleic Acids Research. 27・7 (in press). (1999)

[文献書誌] Ikawa,Y.: "A conserved motif in group IC3 introns is a new class of GNRA receptor" Nucleic Acids Research. 27・8 (in press). (1999)

[文献書誌] Komatsu,Y.: "Modification and application of self-trimming hairpin ribozymes to targeting a transcribed RNA in vitro." J.Biochem. 124・5. 986-991 (1998)

[文献書誌] Koizumi,M.: "Design and anti-HIV-1 activity of hammerhead and hairpin ribozymes containing a stable loop." Nucleosides & Nucleotides. (in press).

[文献書誌] Kobayashi,H: "Effets of a high-affinity antibody fragment on DNA polymerase reactions near a (6-4) photoproduct site." Photochem.& Photobio.69・2. 226-230 (1999)

[文献書誌] Ota,N.: "Determination of interactions between structured nucleic acids by fluorescence resonance energy transfer (FRET): selection of target sites for functional nucleic acids" Nucleic Acids Res.26. 735-743 (1998)

[文献書誌] Kawasaki,H.: "p300 and ATF-2 are components of the DRF complex, which regulates retinoicacid-and E1A-mediated transcription of the c-jun gene in F9 cells." Genes & Dev.12. 233-245 (1998)

[文献書誌] Kawasaki,H.: "Distinct roles of the co-activators p300 and CBP in retinoic-acid-induced F9-cell differentiation." Nature. 393. 284-289 (1998)

[文献書誌] Ota,N.: "Effects of helical structures formed by the binding arms of DNAzymes and their substrates on catalytic activity" Nucleic Acids Res.26. 3385-3391 (1998)

[文献書誌] Kuwabara,T.: "Formation of a catalytically active dimer by tRNA Val-driven short ribozymes" Nature Biotechnology. 16. 961-965 (1998)

[文献書誌] Kuwabara,T.: "A novel nallosterically trans-activated ribozyme, the maxizyme, with exceptional specificity in vitro and in vivo." Mol.Cell. 2. 617-627 (1998)

[文献書誌] Kuwabara,T.: "tRNA Val-heterodimeric maxizymes with high potential as gene-inactivating agent: simultaneous cleavage at two sites in HIV-1tat mRNA in cultured cells." Proc.Natl.Acad.Sci.USA96. 1886-1891 (1999)

[文献書誌] Shibuya,T.: "Characterization of the ptr6+ gene in fission yeast: A possible involvement of a transcriptional coactivator TAF in nucleocytoplasmic transport of mRNA" Genetics. in press (1999)

[文献書誌] Kanai,N.: "A “G" to “A" transition at the last nucleotide of exon 6 of the gene(868G-A) may result in either a splice or missense mutation in patients with X-linked severe combined immunodeficiency" Human Genet.104. 36-42 (1999)

[文献書誌] Habara,Y.: "The fission yeast prp10+ gene involved in pre-mRNA splicing encodes a homologue of highly conserved splicing factor, SAP155" Nucleic Acids Res.26・24. 5662-5669 (1998)

[文献書誌] Imamura,O: "Cloning and characterization of a human DEAH-box RNA helicase, a functional homolog of fission yeast Cdc28/Prp8." Nucleic Acids Res.26. 2063-2068 (1998)

[文献書誌] Nitta,I.: "Possible involvement of Escherichia coli 23S ribosomal RNA in peptide bond formation." RNA. 4. 257-267 (1998)

[文献書誌] Nitta,I.: "Reconstitution of peptide bond formation with Escherichia coli 23S ribosomal RNA domains." Science. 281. 666-669 (1998)

[文献書誌] Nitta,I.: "A novel cell-free system for peptide synthesis driven by pyridine." Biol.Chem.379. 819-829 (1998)

[文献書誌] Kimoto,M.: "RNA aptamers that specifically bind to the Ras-binding domain of Raf-1" FEBS Letters. 441. 322-326 (1998)

[文献書誌] Kiga,D.: "An RNA aptamer to the xanthine/guanine base with a distictive mode of purine recognition" Nucleic Acids Research. 26・7. 1755-1760 (1998)

[文献書誌] Seio,K.: "Chemical synthesis and properties of conformationally fixed diuridine monophosphates as builiding blocks of the RNA turn motif" Journal of Organic Chemistry. 63・5. 1429-1443 (1998)

1998 年度実績報告書

井上丹京都大学, 大学院・理学研究科, 教授 (40114855)