1995 Fiscal Year Final Research Report Summary
Developmental research on the microscopical imaging of DNA transcription process.
| Project/Area Number |
06558101
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| Research Category |
Grant-in-Aid for Developmental Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
Biophysics
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| Research Institution | Nagoya University |
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Principal Investigator |
USUKURA Jiro Nagoya University School of Medicine Associate Professor, 医学部, 助教授 (30143415)
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| Co-Investigator(Kenkyū-buntansha) |
OBATA Shuichi Nagoya University School of Medicine Instructor, 医学部, 助手 (10204273)
WAKABAYASHI Takashi Nagoya University School of Medicine Professor, 医学部, 教授 (00079998)
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| Project Period (FY) |
1994 – 1995
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| Keywords | DNA / Transcription, / RNA Polymerase / Electron microscopy / CRP / Low angle shadowing / Atomic force microscopy ^<5)> |
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| Research Abstract |
This project aimed to establish the imaging methods of transcription process of DNA,especially three dimensionalassembly of DNA,RNA polymerase and regulatory proteins complex. For this purpose, we developed first freeze-drying apparatus by molecular distilllation, that is available for atomic force imaging. Howefver, atomic force microscope was not able to described molecular architecture of DNA-protein complex. rather classical low angle shadowing detected obviously the complex structure. To improve the resolution of shadowing method, we developed new freeze etching machine working at extremely high vacuum (10^<-6> P). Platinum particles evaporated in this apparatus at-100ﾟC was so small that basic structure of DNA as well as molecular organization of DNA binding proteins such as polymerase was depicted by low angle rotary shadowing.therefore, we used this technique exclusively for imaging of DNA transcription processes. High resolution rotary shadowing revealed that E.coli RNA polymerase consisted of 5 subunits which is well consistentwith previous biochemical evidence. Depending upon molecular weight of subunits, structures corresponding to alpha, beta, gamma subunits were identified respectively. alpha, alpha andbeta, beta'subunit formed two channels. One of them, larger one, was located between 2 alphaandbeta, beta'. The another one was found between beta and beta'like a slit. Assembled DNA was always observed in the groove between 2alpha and beta, beta'. These are suggested that newlysynthesized mRNA may come up along the slit formed by betaandbeta'.
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Research Products
(11 results)
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[Publications] Liu, X., Seno, K., Nishizawa, Y., Hayashi, F., Yamazaki, A., Matsumoto, H., Wakabayashi, T.& Usukura, J.: "Ultrastructural localization of retinal guanylate cyclase in human and monkey retinas." Exp.Eye Res.59. 761-768 (1994)
Description
「研究成果報告書概要(欧文)」より
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[Publications] Arai, M., Nishizawa, Y., Liu, X., Usukura, J.Awaya, S.&, Wakabayashi, T.: "Confocal imaging of megamitochondria formation induced by ethanol and hydrazine in culture cells (RL-34)" Bioimages. 3. 25-30 (1995)
Description
「研究成果報告書概要(欧文)」より
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