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Crystallographic Studies of Flavoreductases in Electron Transport Systems of Liver Microsomes

Research Project

Project/Area Number 02680219
Research Category

Grant-in-Aid for General Scientific Research (C)

Allocation TypeSingle-year Grants
Research Field 生物物性学
Research InstitutionTokyo Institute of Technology (1991)
Osaka University (1990)

Principal Investigator

MIKI Kunio  Tokyo Inst. of Tech. Res. Lab. of Resources Utilization Associate Professor, 資源化学研究所, 助教授 (10116105)

Project Period (FY) 1990 – 1991
Project Status Completed (Fiscal Year 1991)
Budget Amount *help
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1991: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1990: ¥1,200,000 (Direct Cost: ¥1,200,000)
KeywordsX-ray crystal structure analysis / Three-dimensional structure / Flavoprotein / Reductases / Electron transport / X形結晶解析 / フラビン
Research Abstract

We have carried out X-ray crystal structure analyses of two flavoreductases (NADH-cytochrome b_5 reductase and HADPH-cytochrome P450 reductase) located in two electron transport pathways in liver microsomes where NADH and HADPH function as initial electron donors, respectively.
NADH-cytochrome b_5 reductase has so far been crystallized by ourselves. We searched two good heavy-atom derivatives in order to obtain the electron density map based on the multiple isomorphous replacement procedure. In spite of many attempts, however, we could find only one derivative. Therefore, we calculated the electron density map at 5A resolution by the single isomorphous replacement. This electron density map was in good quality which enabled us to distinguish between the protein and solvent regions. Furthermore, we collected intensity data at the higher resolutions and used synchrotron radiation to measure precise intensities including anomalous dispersion effects. We also drawn the electron density map based on these data. We tried to interpret these maps, and then we estimated the outline of the molecular shape and roughly traced the folding of this enzyme. We also estimated the FAD binding position which is one of the most important key points in this projects. However, to make more precise discussion, we need better quality electron density maps. It is the most important problem in the near future to improve the quality of these density maps.
We have tried to crystallize HADPH-cytochrome P450 reductase. Initially we searched precipitants effective to this enzyme and were successful in finding a few reagents. We tried many crystallization conditions using these precipitants. But, we could not yet obtained any good crystals suitable for X-ray diffraction works.

Report

(3 results)
  • 1991 Annual Research Report   Final Research Report Summary
  • 1990 Annual Research Report
  • Research Products

    (8 results)

All Other

All Publications (8 results)

  • [Publications] 藤井 功他: "イメ-ジングプレ-トの生物結晶学への応用ー回折デ-タの一貫処理システム" 日本結晶学会誌. 32. 261-267 (1990)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] I.Fujii他: "Evaluation of Xーray Difraction Data from Protein Crytals with of an Imaging Plate" Acta Crystallographica. B47. 137-144 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] 三木 邦夫他(共著)(分担): "新生化学実験講座、第1巻「タンパク質」III.高次構造" 東京化学同人, 436 (1990)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] 三木 邦夫他(共著)(分担): "バイオ高分子研究法5 「バイオ高分子研究における物理化学計測とその応用」" 学会出版センタ-, (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] I. Fujii, et. al.: "Evaluation of X-ray Diffraction Data from Protein Crystals with Use of an Imaging Plate" Acta Crystallogr.B47. 137-144 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] 藤井 功 他: "イメ-ジングプレ-トの生物結晶学への応用ー回折デ-タの一貫処理システム" 日本結晶学会誌. 32. 261-267 (1990)

    • Related Report
      1991 Annual Research Report
  • [Publications] I.Fujii 他: "Evaluation of Xーray Diftraction Data from Protein Crystals with Use of an Imaging Plate" Acta Crystalloglaplrica. B47. 137-144 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] 三木 邦夫 他(共著): "バイオ・高分子研究法5「バイオ高分子研究における物理化学計測とその応用」" 学会出版センタ-, (1992)

    • Related Report
      1991 Annual Research Report

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Published: 1990-04-01   Modified: 2016-04-21  

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