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Preparation of crystals of cytochrome c oxidase and cytochrome bcl complex

Research Project

Project/Area Number 06453220
Research Category

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

Allocation TypeSingle-year Grants
Research Field Biophysics
Research InstitutionHimeji Institute of Technology

Principal Investigator

YOSHIKAWA Shinya  Himeji Institute of Technology, Department of Life Science, Professor, 理学部, 教授 (40068119)

Project Period (FY) 1994 – 1995
Project Status Completed (Fiscal Year 1995)
Budget Amount *help
¥7,100,000 (Direct Cost: ¥7,100,000)
Fiscal Year 1995: ¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 1994: ¥4,800,000 (Direct Cost: ¥4,800,000)
KeywordsMembrane proteins / Protein crystallization / X-ray crystallographic analysis / Cytochrome c oxidase / Cytochrome bcl complex / Detergent / チトクロムbc1複合体 / 結晶化
Research Abstract

Effect of detergent structure on crystallization of bovine heart cytochrome c oxidase has been examined extensively. Decylmaltoside provides crystals which diffract x-rays up to 2.2 A^^゚ resolution or to better resolution, although at fairly low reproducibility A slight modification in the detergent structure affects the crystallization conditions significantly. For example, neither nonyl nor undecyl maltosides provides stable crystals. Structure of the sugar moiety in decylmaltoside is critical for the crystallization. Actually, no sugar other than maltose provides even microcrystals of bovine heart cytochrome oxidase.
Crystallization conditions for various derivalives of this enzyme stabilized with decylmaltoside has been established. Crystals of azide bound form as well as cyanide bound form are prepared from soaking the crystals of fully oxidized form in the medium containing each ligands. Crystals of the fully reduced form and the fully reduced CO bound form was not able to be stab … More ilized under nitrogen atomosphere, because of the strong sensitivity to O2. However, it is impossible to fix a crystal in a capillary in the presence of mother lizuer. Thus, the crystals of the fully reduced form and of the fully reduced CO bound form are stabilized by sephadex gel equilibrated with the mother liquer containing ascorbate and cytochrome c as the reducing system and saturated with N2 and CO respectively. The oxidation and ligand binding states are checked by the absorption spectra of the crystals placed in the mother liquer without sephadex gel. The crystals of fully reduced form diffracts x-rays up to 2.2 A^^゚ resolution and the other derivative crystals diffract to better than 2.9 A^^゚ resolution. Cytochrome bcl complex stabilized with sucrose monocaplate provides crystals which diffract to 2.8 A^^゚ resolution. However this crystallization conditions are not so sensitive to the structure of detergents as in the case of bovine heart cytochrome c oxidase. Purity of deoxycholate which is for solubilizing the cytochrome bcl complex from mitochondrial inner membrane seems critical for the crystallization. Less

Report

(3 results)
  • 1995 Annual Research Report   Final Research Report Summary
  • 1994 Annual Research Report
  • Research Products

    (36 results)

All Other

All Publications (36 results)

  • [Publications] M.Kawamoto: "New crystal forms and preliminary X-ray diffraction studies of mitochondrial cytochrome bc_1 complex from bovine heart." J.Mol.Biol.244. 238-241 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] K.Shinzawa-Itoh: "Effects of ethyleneglycol chain length of dodecyl polyethyleneglycol monoether on the crystallization of bovine heart cytochrome c oxidase." J.Mol.Biol.247. 572-575 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] T.Tsukihara: "Structures of metal sites of oxidized bovine heart cytochrome c oxidase at 2.8Å." Science. 269. 1069-1074 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] T.Tsukihara: "The whole structure of the 13 subunit oxidized bovine heart cytochrome c oxidase at 2.8Å." Science. 272. 1136-1144 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Yoshikawa: "Crystalstructure of fully oxidized bovine heart ccytochrome c oxidase at 2.8 A resolution: a review." Biochemistry(Moscow). 61. 1369-1376 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Murakami: "Crystals of bovine heart ubiquinol-cytochrome c reductase diffracting X-rays up to 2.8 A resolution at 3 C." Acta Cryst.D54. 146-147 (1998)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Hirota, T.Ogura, Shinzawa-Itoh, S.Yoshikawa, M.Nagai and T.Kitagawa: "Vibrational assignments of the FeCO unit of CO-bound heme proteins revisited : Observation of a new CO-isotope-sensitive Raman band assignable to the FeCO bending fundamental" J.Phys.Chem. 98. 6652-6660 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] M.Kawamoto, T.Kubota, T.Matsunaga, K.Fukuyama, H.Matsubara, K.Shinzawa-Itoh and S.Yoshikawa: "New crystal forms and preliminary X-ray diffraction studies of mitochondrial cytochrome bcl complex from bovine heart" J.Mol.Biol. 244. 238-241 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Hirota, T.Ogura, E.H.Appelman, K.Shinzawa-Itoh, S.Yoshikawa and T.Kitagawa: "Observation of a new oxygenisotope-sensitive Raman band for oxyhemoproteins and its implications in heme pocket structure" J.Am.Chem.Soc.116. 10564-10570 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] D.A.Proshlyakov, T.Ogura, K.Shinzawa-Itoh, S.Yoshikawa, E.A.Appelman and T.Kitagawa: "Selective resonance Raman observation of the "607 nm" form generated in the reaction of oxidized cytochrome c oxidase with hydrogen peroxide" J.Biol.Chem. 269. 29385-29388 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] K.Shinzawa-Itoh, H.Ueda, S.Yoshikawa, H.Aoyama, E.Yamashita and T.Tsukihara: "Effects of ethyleneglycol chainlength of dodecyl polyethyleneglycol monoether on the crystallization of bovine heart cytochrome c oxidase" J.Mol.Biol. 247. 572-575 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] T.Tsukihara, H.Aoyama, E.Yamashita, T.Tomizaki, H.Yamaguchi, K.Shinzawa-Itoh, R.Nakashima, R.Yaono and S.Yoshikawa: "Structures of metal sites of oxidized bovine heart cytochrome c oxidase at 2.8A^^゚" Science. 269. 1069-1074 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Yoshikawa, M.Mochizuki, X.-J.Zhao, and W.S.Caugkey: "Effects of overall oxidation sate on infrared spectra of heme a3 cyanide in bovine heart cytochrome c oxidase" J.Biol.Chem. 270. 4270-4279 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] T.Tsukihara, K.Aoyama, E.Yamashita, T.Tomizaki, H.Yamaguchi, K.Shinzawa-Itoh, R.Nakashima, R.Yaono and S.Yoshikawa: "The whole structure of the 13 subunit oxidized bovine heart cytochrome c oxidase at 2.8A^^゚" Science. 272. 1136-1144 (1966)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] D.A.Proshlyakov, T.Ogura, K.Shinzawa-Itoh, S.Yoshikawa and T.Kitagawa: "Resonance Raman/absorption characterization of the oxo intermediates of cytochrome c oxidase generated in its reaction with hydrogen peroxide : pH and H202 concentration dependence" Biochemistry. 35. 8580-8586 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Hirota, T.Ogura, K.Shinzawa-Itoh, S.Yoshikawa, and T.Kitagawa: "Observation of multiple CN-isotope-sensitive Raman bands for CN-adducts of hemoglobin, myoglobin, and cytochrome c oxidase : evidence for vibrational coupling berween the Fe-C-N bending and porphyrin in-plane modes" J.Phys.Chem.37. 15274-15279 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] T.Ogura, S.Hirota, D.A.Proshlyakov, K.Shinzawa-Itoh, S.Yoshikawa and T.Kitagawa: "Time resolved resonance Reman evidence for tight coupling between electron transfer and proton pumping of cytochrome c oxidase upon the change from the Fe^V oxidation level to the Fe^<IV> oxidation level" J.Am.Chem.Soc. 118. 5443-5449 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Yoshikawa, T.Tsukihara and K.Shinzawa-Itoh: "Crystalstructure of fully oxidized bovine heart ccytochrome c oxidase at 2.8 Aresolution : a review" Biochemistry (Moscow). 1369-1376 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Yoshikawa: "Beef heat cytochrome c oxidase" Curr.Opin.Struct.Biol.7. 574-579 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Murakami, K.Shinzawa-Itoh, S.Yoshikawa: "Crystals of bovine heart ubiquinol-cytochrome c reductase diffracting X-rays upto 2.8 A resolution at 3 C" Acta Cryst.Ser. D54. 146-147 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] E.Yamashita, H.Yamaguchi, T.Tomizaki, H.Aoyama, T.Tsukihara, K.Shinzawa-Itoh, R.Nakashima, R.Yaono and S.Yoshikawa: "Crystal structure analysis of bovine heart cytochrome c oxidase at 2.8 A resolution" Acta Cryst.Ser.D. in press. (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Yoshikawa, K.Shinzawa-Itoh, R.Nakashima, R,Yaono, E.Yamashita, N.Inoue, M.Yao, M.J.Fei, C.Peters Libeu, T.Mizushima, H.Yamaguchi, T.Tomizaki and T.Tsukihara: "Redox coupled crystal structural changes in bovine heart cytochrome c oxidase" Science. submitted. (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] S.Yoshikawa, K.Shinzawa-Itoh and T.Tsukihara: Crystal structure and reaction mechanism of bovine heart cytochrome c oxidase, in Oxygen Homeostasis and Its Dynamics, Keio University Symposia for Life Science and Medicine (Y.Ishimura, H.Shimada and M.Suematsu eds.). Springer-Verlag, Tokyo, 13-23 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] T.Kitagawa, T.Ogura, S.Hirota, D.A.Proshlyakov, J.Matysik, E.H.Appelman, K.Shinzawa-Itoh and S.Yoshikawa: Time-resolved resonance Raman study of dioxygen reduction by cytochrome c oxidase, in Oxygen Homeostasis and Its Dynamics, Keio University Symposia for Life Science and Medicine (Y.Ishimura, H.Shimada and M.Suematsu eds.). Springer-Verlag, Tokyo, 57-71 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] T.Tsukihara,et al.: "The whole structure of the 13 subunit oxidized bovine heart cytochrome c oxidase at 2.8 Å。" Science. in press。 (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] D.A.Proshlyakov.et al.: "Microcirculating system for simultaneous determination of Raman and absorption spectra of enzymatic reaction intermediates and its application to the reaction" Biochemistry. 35. 76-82 (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] T.Tsukihara,et al.: "Structures of metal sites of oxidized bovine heart cytochrome c oxidase at 2.8 Å." Science. 269. 1069-1074 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] S.Yoshikawa,et al.: "Effects of overall oxidation state on infrared spectra of Heme a3 cyanide in bovine heart cytochrome c oxidase." J.Biol.Chem.270. 4270-4279 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] K.Shinzawa-Itoh,et al.: "Effects of ethyleneglycol chain length of dodecyl polyethyleneglycol monoether on the srystallization of bovine heart cytochrome c oxidase." J.Mol.Biol.247. 572-575 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] M.Kawamoto,et al.: "New crystal forms and preliminary X-ray difffraction studies of mitochondrial bc1 complex from bovine heart." J.Mol.Biol.244. 238-241 (1994)

    • Related Report
      1995 Annual Research Report
  • [Publications] S.Yoshikawa: "Effects of overall oxidatio state on infrared spectra of Heme a_3 cyanide in bovine heart cytochrome c oxidase" J.Biol.Chem.270. 4270-4279 (1995)

    • Related Report
      1994 Annual Research Report
  • [Publications] K.Shinzawa-Itoh: "Effects of ethy leneglycol chain length of dodecyl polyethyleneglycol monoether on the crystallization of bovine heart cytochrome c oxidase" J.Mol.Biol.246. 572-575 (1995)

    • Related Report
      1994 Annual Research Report
  • [Publications] D.A.Proshlyakov: "Selective resonance Raman observation of the “607 nm"form generated in the reaction of oxidized cytochrome c oxidase with hydrogen peroxide" J.Biol.Chem.269. 29385-29388 (1994)

    • Related Report
      1994 Annual Research Report
  • [Publications] S.Hirota: "Observation of a new oxygen-isotope-sensitive Raman band for oxyhemoproteins and its implications in heme pocket structure" J.Am.Chem.Soc.116. 10564-10570 (1994)

    • Related Report
      1994 Annual Research Report
  • [Publications] M.Kawamoto: "New crystal forms and preliminary X-ray deffraction studies of mitochondrial cytochrome bc_1 complex from bovine heart" J.Mol.Biol.244. 238-241 (1994)

    • Related Report
      1994 Annual Research Report
  • [Publications] S.Hirota: "Vibrational assignments of the FeCO unit of CO-bound heme proteins revisited:observation of a new CO-isotope-sensitive raman band assignable to the FeCO bending fundamental" J.Phys.Chem.98. 6652-6660 (1994)

    • Related Report
      1994 Annual Research Report

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Published: 1994-04-01   Modified: 2016-04-21  

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