1990 Fiscal Year Annual Research Report

複製開始域におけるDNAと蛋白の相互作用ー大腸菌染色体の複製、分配、細胞分裂の分子的基礎

Research Project

Project/Area Number	02404088
Research Institution	National Institute of Genetics
Principal Investigator	堀内賢介国立遺伝学研究所, 細胞遺伝研究系, 教授 (70219210)
Co-Investigator(Kenkyū-buntansha)	西村昭子国立遺伝学研究所, 微生物保存研究室, 助手 (20142002) 東谷篤志国立遺伝学研究所, 細胞遺伝研究系, 助手 (40212162) 原弘志国立遺伝学研究所, 細胞遺伝研究系, 助手 (00173071) 西村行進国立遺伝学研究所, 細胞遺伝研究系, 助手 (30029699) 安田成一国立遺伝学研究所, 細胞遺伝研究系, 助教授 (90037250)
Keywords	DNA複製 / 繊維状ファ-ジ / 大腸菌 / 超螺旋 / ペニシリン結合蛋白 / 細胞分裂 / fts遺伝子群 / 染色体地図
Research Abstract	1.大腸菌繊維状ファ-ジのプラス鎖複製開始域における、複製開始蛋白gp2及びIHF蛋白のDNAとの反応を研究した結果、(1)gp2のnicking活性とclosing活性の比が基質DNAの負超螺旋度に依存すること、(2)IHF非依存性変異を持つgp2のnicking活性は、負超螺旋度に殆ど依らない点で野生型と著しく異なることを発見した。これはIHFが負超螺旋と類似の機構によってDNA複製開始反応にあずかることを示唆する。2.同ファ-ジのマイナス鎖複製開始域において多数の欠失及び挿入変異を分離し、そのorigin活性をin vivo及びin vitroで解析した。その結果、RNA primer合成開始点を含む十数塩基の欠失はマイナス鎖合成に強い影響がないこと、その下流の約110塩基がorigin活性にとって重要であることを明らかにした。更に、マイナス鎖複製欠損変異ファ-ジは感染菌においてSOS機能を誘発することを発見した。これは菌体内においてSOSの誘発が単鎖DNAそのものに依り、複製フォ-ク等の構造を要しないことを示唆する。3.大腸菌染色体分配に関わるparC及びparEが、新しいtopoisomerase(topoIV)をcodeすることを見出した(東大・医科研・加藤潤一氏と共同)。4.大腸菌<prc>___ー遺伝子は細胞分裂隔壁形成酵素であるペニシリン結合蛋白質3の前駆体C末端のprocessingにあずかる。<prc>___ー^ー株ではペリプラズム蛋白の培地中への漏出が認められるが、これが<pld>___ーA変異(細胞外膜のフォスフォリパ-ゼAの欠損)によって抑圧されることを見出した。5.細菌細胞壁のペプチドグリカン合成はバクトプレニル燐酸分子上で起こるが、その前駆体であるファルネシル二燐酸の合成酵素の遺伝子<isp>___ーAを同定し、その全塩基配列を決定した(岐阜大・藤崎真吾氏と共同)。6.pLCーplasmid collectionによる相補テスト、及びpLCーplasmidと小原整列libraryとのhybridizationにより、細胞分裂に関与する356のfts変異のmappingを完成した。

Research Products

(16 results)

All Other

All Publications (16 results)

[Publications] 堀内賢介: "繊維状ファ-ジのDNA複製開始域" 遺伝学雑誌. 65. 225-241 (1990)
[Publications] Nishimura,Akiko: "A rapid and highly efficient method for preparation of competent Escherichia coli cells." Nucleic Acids Res.18. 6169-6169 (1990)
[Publications] Kato,Junーichi: "New topoisomerase essential for chromosome segregation in E.coli." Cell. 63. 393-404 (1990)
[Publications] Fujisaki,Shingo: "Cloning and nucleotide sequence of the ispA gene responsible for fornesyl diphosphate synthase activity in Escherichia coli." J.Biochem.108. 995-1000 (1990)
[Publications] Greenstein,David: "Replicationーindependent mutation increases the coーoperativity with which an initiator protein binds its origin." J.Mol.Biol.211. 91-101 (1990)
[Publications] Higashitani,Atsushi: "Replication of filamentous phage fl:Enhancerーindependent mutation of an initiator protein (gpII)." Annual Report of National Institute of Genetics. 40. 43-44 (1990)
[Publications] Higashitani,Atsushi: "Multicopy suppression in E.coli:Cloning of sufB,a dosagedependent suppressor of a thermosensitive allーdivision mutation,ftsB." Annual Report of National Institute of Genetics. 40. 44-46 (1990)
[Publications] Kato,Junーichi: "A novel topoisomerase essential for the E.coli chromosome partition." Annual Report of National Institute of Genetics. 40. 46-48 (1990)
[Publications] Hara,Hiroshi: "Identificatior and cellular localization of the product of prc,a gene involved in the Cーterminal processing of penicillinーbinding protein 3 of Escherichia coli." Annual Report of National Institute of Genetics. 40. 49-51 (1990)
[Publications] Yamamoto,Yoshihiro: "Nucleotide sequence of the prc gene of Escherichia coli." Annual Report of National Institute of Genetics. 40. 51-52 (1990)
[Publications] Fujisaki,Shingo: "A mutant producing temperatureーsensitive fornesyl diphosphate synthase of Escherichia coli." Annual Report of National Institute of Genetics. 40. 52-53 (1990)
[Publications] Nishimura,Akiko: "Mapping a whole set of cell division genes in Escherichia coli kー12." Annual Report of National Institute of Genetics. 40. 53-54 (1990)
[Publications] Nishimura,Akiko: "Correlation of pLCーplasmids to the physical map of Escherichia coli Kー12."
[Publications] Hara,Hiroshi: "Cloning,mapping and characterization of the Escherichia coli prc gene involved in Cーterminal processing of penicillinーbinding protein 3."
[Publications] Higashitani,Atsushi: "Replication enhancerーindependent mutations reduce superhelicity requirement of the initiator protein of bacteriophage f1."
[Publications] Nishimura,Akiko: "Control of Cell Growth and Division (H.Yoshikawa and A.Ishihama,eds.)の内、Mapping of a whole set of cell division genes in Escherichia coli Kー12." Academic Press, (1991)

1990 Fiscal Year Annual Research Report

複製開始域におけるDNAと蛋白の相互作用ー大腸菌染色体の複製、分配、細胞分裂の分子的基礎

Principal Investigator

堀内 賢介 国立遺伝学研究所, 細胞遺伝研究系, 教授 (70219210)

Research Products

[Publications] 堀内 賢介: "繊維状ファ-ジのDNA複製開始域" 遺伝学雑誌. 65. 225-241 (1990)

[Publications] Nishimura,Akiko: "A rapid and highly efficient method for preparation of competent Escherichia coli cells." Nucleic Acids Res.18. 6169-6169 (1990)

[Publications] Kato,Junーichi: "New topoisomerase essential for chromosome segregation in E.coli." Cell. 63. 393-404 (1990)

[Publications] Fujisaki,Shingo: "Cloning and nucleotide sequence of the ispA gene responsible for fornesyl diphosphate synthase activity in Escherichia coli." J.Biochem.108. 995-1000 (1990)

[Publications] Greenstein,David: "Replicationーindependent mutation increases the coーoperativity with which an initiator protein binds its origin." J.Mol.Biol.211. 91-101 (1990)

[Publications] Higashitani,Atsushi: "Replication of filamentous phage fl:Enhancerーindependent mutation of an initiator protein (gpII)." Annual Report of National Institute of Genetics. 40. 43-44 (1990)

[Publications] Higashitani,Atsushi: "Multicopy suppression in E.coli:Cloning of sufB,a dosagedependent suppressor of a thermosensitive allーdivision mutation,ftsB." Annual Report of National Institute of Genetics. 40. 44-46 (1990)

[Publications] Kato,Junーichi: "A novel topoisomerase essential for the E.coli chromosome partition." Annual Report of National Institute of Genetics. 40. 46-48 (1990)

[Publications] Hara,Hiroshi: "Identificatior and cellular localization of the product of prc,a gene involved in the Cーterminal processing of penicillinーbinding protein 3 of Escherichia coli." Annual Report of National Institute of Genetics. 40. 49-51 (1990)

[Publications] Yamamoto,Yoshihiro: "Nucleotide sequence of the prc gene of Escherichia coli." Annual Report of National Institute of Genetics. 40. 51-52 (1990)

[Publications] Fujisaki,Shingo: "A mutant producing temperatureーsensitive fornesyl diphosphate synthase of Escherichia coli." Annual Report of National Institute of Genetics. 40. 52-53 (1990)

[Publications] Nishimura,Akiko: "Mapping a whole set of cell division genes in Escherichia coli kー12." Annual Report of National Institute of Genetics. 40. 53-54 (1990)

[Publications] Nishimura,Akiko: "Correlation of pLCーplasmids to the physical map of Escherichia coli Kー12."

[Publications] Hara,Hiroshi: "Cloning,mapping and characterization of the Escherichia coli prc gene involved in Cーterminal processing of penicillinーbinding protein 3."

[Publications] Higashitani,Atsushi: "Replication enhancerーindependent mutations reduce superhelicity requirement of the initiator protein of bacteriophage f1."

[Publications] Nishimura,Akiko: "Control of Cell Growth and Division (H.Yoshikawa and A.Ishihama,eds.)の内、Mapping of a whole set of cell division genes in Escherichia coli Kー12." Academic Press, (1991)

堀内賢介国立遺伝学研究所, 細胞遺伝研究系, 教授 (70219210)

[Publications] 堀内賢介: "繊維状ファ-ジのDNA複製開始域" 遺伝学雑誌. 65. 225-241 (1990)