| Project/Area Number |
63580205
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
分子遺伝学・分子生理学
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
WADA Chieko Kyoto University, Institute for Virus Research, Instructor, ウィルス研究所, 助手 (10175698)
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| Co-Investigator(Kenkyū-buntansha) |
YURA Takashi Kyoto University, Institute for Virus Research, Professor, ウィルス研究所, 教授 (20027311)
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| Project Period (FY) |
1988 – 1990
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| Project Status |
Completed (Fiscal Year 1989)
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| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1989: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1988: ¥1,600,000 (Direct Cost: ¥1,600,000)
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| Keywords | Replication initiation Of mini-F DNA / Transcription of repE gene / sigma 32 factor / Dnak protein / DnaJ protein / GrpE protein / Copy mutant of mini-F / ミニFコピ-変異 / 大腸菌Fプラスミド / RepE蛋白質 / σ^<32>RNAポリメラ-ゼ / FrepEプロモ-タ- / PlrepAプロモ-タ- / Rtsl repAプロモ-タ- / 太腸菌Fプラスミド / repEプロモーター / σ^<32>RNAポリメラーゼ / Rtsl |
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| Research Abstract |
Replication of F, mini-F and related plasmids ( mini-Rts1, mini-P1 ) is stringently controlled to maintain their copy number of 1 to 2 per host chromosome. It is thought that the plasmid copy number is regulated primarily by the amount or activity of the specific initiator protein essential for replication of the respective plasmid. We have found that the mini-F repE gene encoding the initiator protein is transcribed by RNA polymerase containing delta^<32> and that mini-F can not replicate in DELTA rpoH cells lacking delta^<32> protein. A subset of E.coli heat shock protein DnaK, DnaJ and GrpE are known to be required for lambda phage DNA replication and for growth of E.coli cells probably at all temperature. We have shown that these heat shock proteins are also required for replication of mini-F plasmid. When excess amounts of the replication initiator protein (RepE) were provided by means of a multicopy plasmid carrying repE,mini-F can replicate in the condition without DnaK, J, GrpE protein. Furthermore, we have isolated and characterized mini-F mutants able to replicate in the absence of delta^<32>. These mini-F plasmid produced altered initiator protein and exhibited a very high copy and were able to replicate in strains deficient in any of the above heat shock proteins. These results indicate that the subset of heat shock proteins play essential roles that help the functioning of the RepE initiator protein in mini-F DNA replication. We found that mini-Rts1 and mini-P1 like mini-F cannot replicate in the rpoH strain. The rep genes of both these plasmids like the mini-F were found to be transcribed by RNA polymerase delta^<32> as well as by delta^<70>. The replication of mini-Rts1 and mini-P1 like mini-F also depends on DnaK,DnaJ and GrpE protein.
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