| Project/Area Number |
03454032
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
発酵工学
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| Research Institution | HIROSHIMA UNIVERSITY |
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Principal Investigator |
NIMI Osamu Hiroshima Univ., Fac. Engin., Professor, 工学部, 教授 (60034360)
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| Co-Investigator(Kenkyū-buntansha) |
SHINKAWA Hidenori Hiroshima Univ., Fac. Engin., Research Associate, 工学部, 助手 (50226338)
KINASHI Haruyasu Hiroshima Univ., Fac. Engin., Associate Professor, 工学部, 助教授 (80224997)
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| Project Period (FY) |
1991 – 1992
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| Project Status |
Completed (Fiscal Year 1992)
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| Budget Amount *help |
¥5,900,000 (Direct Cost: ¥5,900,000)
Fiscal Year 1992: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1991: ¥4,000,000 (Direct Cost: ¥4,000,000)
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| Keywords | Streptomyces / Pleiotropic Mutation / Streptomyces griseus / Sigma Factor / hrdB / RNA Polymerase / プロモ-タ-結合蛋白 |
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| Research Abstract |
In order to clue a control mechanism of gene expression in secondary metabolism, RNA polymerase activity and sigma factor were determined in pleiotropic mutant derived from Streptomyces griseus. Results obtained are as follow
1. RNA polymerase were purified in both parent and pleiotropic mutant. In vitro transcriptional activities using both principal and secondary metabolic gene as template were determined. RNA polymerase from mutant possessed no activity transcribing secondary metabolic gene, suggesting that a sigma factor was unusual or lack in the mutant.
2. A protein binding to promoter region on strB gene which contributed to streptomycin biosynthesis was screened. A binding protein attached to promoter gene was foud, but the protein was non specific for pleiotropic mutant.
3. A sigma factor in S. griseus was screened. Gene hrdB of S. coelicolor A3(2) was used as probe for both colony and Southern hybridization experiment. A clone houvouring pCE8 was hybridized strongly to the probe DNA.
4. Nucleotide sequences of pCE8 was determined. A ORF(1335bp) was found in the sequence. The ORF contained 36bp rpoB box, which was preserved nucleotide sequence in gene of several sigma factor. DNA homology of hrdB to pCE8 was 86.3%.
5. The sigma like protein derived from pCE8 were determined in several phase of growth in streptomyces mycelium, using Northern bloting method. The protein was recognized in all phases in myceliul growth. These result revealed that the protein from pCE8 was not secondary metabolic factor but principal one and suggested the presence of another sigma factor concerning secondary metabolism in Streptomyces.
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