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Studies on active anion transport systems in a biological membrane

Research Project

Project/Area Number 04044123
Research Category

Grant-in-Aid for international Scientific Research

Allocation TypeSingle-year Grants
SectionJoint Research
Research InstitutionOkayama University

Principal Investigator

OHMORI Shinji  Faculty of Pharmaceutical Sciences, Okayama University, 薬学部, 教授 (10032872)

Co-Investigator(Kenkyū-buntansha) SHIOZAWA Jud  マックス, プランク生化学研究所, 研究員
LOTTSPEICH Friedrich  Max-Planck-Institute of Biochemistry, プランク生化学研究所, 教授
OESTERHELT Dieter  Max-Planck-Institute of Biochemistry, プランク生化学研究所・所長, 教授
MORITANI Chie  Faculty of Pharmaceutical Sciences, Okayama University, 薬学部, 助手 (60253001)
OTSUKA Masato  Faculty of Pharmaceutical Sciences, Okayama University, 薬学部, 助手 (30243489)
IKEDA Mikiko  Faculty of Health and Welfare Science, Okayama Prefectural University, 保健福祉学部, 教授 (20112154)
SHIOZAWA Judith a.  Max-Planck-Institute of Biochemistry
JUDITH A. Sh  マックス, プランク生化学研究所・膜生化学部門, 研究員
FRIEDRICH Lo  マックス, プランク生化学研究所・遺伝子センター, 教授
DIETER Oeste  マックス, プランク生化学研究所・膜生化学部門, 教授
Project Period (FY) 1992 – 1993
Project Status Completed (Fiscal Year 1993)
Budget Amount *help
¥5,500,000 (Direct Cost: ¥5,500,000)
Fiscal Year 1993: ¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 1992: ¥2,700,000 (Direct Cost: ¥2,700,000)
KeywordsCl^--ATPase / cloning / F type ATPase / sulfate permease / cysA gene / sbp gene / ABC type ATPase / Acetabularia acetabulum / Cl^-ATPase / アニオン / 一次性能動輸送系 / 遺伝子クローニング
Research Abstract

1. Cl^--translocating ATPase
(1) Molecular cloning of the b (50 kDa) subunit : Overlapping 4 clones, pCLBl to 4 were cloned and sequenced by RT-PCR of total RNA and poly(A)^+ RNA isolated from Acetabularia acetabulum. Combined clones encoded the entire ORF of the Cl^--ATPase, b subunit which consisted of 478 amino acids and the molecular weight was calculated to be 51,513. The primary structure showed the highest similarity to Chlamydomonas CF_1-ATPase, beta subunit among cation-translocating ATPases, F, P and V(A) type.
(2) Molecular cloning of the a (54 kDa) subunit : A clone, pCLAl (803 bp) encoding the N-terminal half region of the a aubunit gene was obtained and sequenced. The deduced amino acid sequence was ca. 75% identical with that of spinach CF_1-ATPase, alpha subunit.
(3) Northern analysis : We have also obtained putative gene fragments coding for A. acetabulum CF_1- (273 bp) and MF_1-ATPases (332 bp), beta subunits. Northern analysis was performed for total RNA using the three … More beta like gene fragments. Ca. over 10 kb, 7.5 and 3.8 kb of RNA were detected with the probes derived from the Cl^--b subunit. Ca.2.2 kb of RNA positively hybridized with the CF_1- and MF_1-beta specific probes, respectively.
A gene fragment (260 bp) encoding a putative CF_1-alpha subunit was also obtained. Northern analysis demonstrated the expression of different sizes of RNA (ca. 3.2 kb for the Cl^--a and 2.4 kb for the CF_1-alpha).
In conclusion, the Cl^--ATPase belongs to the F type ATPase family and the presence of a small multigene family for the F type ATPase was suggested in this organism.
2. Sulfate permease
(1) Sulfate uptake by sulfate-starved cells : About 3- to 4-fold increase in sulfate uptake was observed for sulfate-starved cells (3 to 4 cm in length).
(2) Molecular cloning : A putative cysA (359 bp) and sbp (552 bp) gene fragments were obtained from cDNA library and total RNA, respectively.
(3) Northern analysis : Ca. 2.4 kb for cysA and 1.55 kb for sbp were detected.
3. Maltose permease : A putative malk gene fragment (369 bp) was also cloned and sequenced. Ca. 1.65 kb of RNA was detected by Northern analysis.
The results of 2 and 3 supported the presence of ATP-binding cassette (ABC) type ATPase in A. acetabulum. This is the first report describing the ATPase specific for bacteria in plant materials. Less

Report

(2 results)
  • 1993 Final Research Report Summary
  • 1992 Annual Research Report
  • Research Products

    (18 results)

All Other

All Publications (18 results)

  • [Publications] Ohhashi,T.,Katsu,T.& Ikeda,M.: "Improvement of reconstitution of the Cl^--translocating ATPase isolated from Acetabularia acetabulum into liposomes and several anion pump characteristics" Biochim.Biophys.Acta. 1106. 165-170 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 池田己喜子,守谷智恵,大森晋爾: "生体膜に存在するアニオン輸送系の研究-Cl輸送性ATPase及び硫酸イオン輸送系の解析" 日本学術振興会「学術月報」. 46. 838-845 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Satoh,S..,Moritani,C.,Ohmori,S.& Ikeda,M.: "Partial purification and characterization of chloroplast ATPase in Acetabularia acetabulum" Biosci.Biotech.Biochem.58(in press). (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 池田己喜子: "細胞におけるアニオンの役割 カサノリに存在するCl輸送性ATPase及び硫酸イオン輸送系の解析" 薬学雑誌. (発表予定). (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Moritani,C.,Rahman,H.,Md.,Ohmori,S.,Yamauchi,M.,de Groot,E.J.,Oesterhelt,D.& Ikeda,M.: "Sulfate uptake system in Acetabularia acetabulum" Planta. (in press). (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Moritani,C.,Ohhashi,T.,Tagaya,M.,Fukui,T.,Lottspeich,F.,Oesterhelt,D.& Ikeda,M.: "Primary structure of the Cl^--translocating ATPase,b subunit of Acetabularia acetabulum" Planta. (in press). (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Ohhashi,T., Katsu,T. & Ikeda,M.: "Improvement of reconstitution of the Cl^--translocating ATPase isolated from Acetabularia acetabulum into liposomes and several anion pump characteristics" Biochim. Biophys.Acta. 1106. 165-170 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Ikeda,M., Moritani,C. & Ohmori,S.: "Studies on anion transport systems in a biological membrane-Cl^--translocating ATPase and sulfate uptake system" Japanese Science Promotion Society, Gakujutu Geppou. 46. 838-845 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Satoh,S., Moritani,C., Ohmori,S. & Ikeda,M.: "Partial purification and characterization of chloroplast ATPase in Acetabularia acetabulum" Biosci.Biotech.Biochem.58(in press). (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Ikeda, M.: "Roles of anions in cells : Studies on the Cl^--translocating ATPase and sulfate uptake system in Acetabularia acetabulum" Yakugaku Zasshi. (in press). (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Moritani,C., Rahman,H., Md., Ohmori,S., Yamauchi,M., de Groot,E.J., Oesterhelt,D. & Ikeda,M.: "Sulfate uptake system in Acetabularia acetabulum" Planta. (in press). (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Moritani,C., Ohhashi,T., Tagaya,M., Fukui,T., Lottspeich,F., Oesterhelt,D. & Ikeda,M.: "Primary structure of the Cl^--translocating ATPase, b subunit of Acetabularia acetabulum" Planta. (in press). (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 池田 己喜子: "生体膜に存在するアニオン輸送系の解明-Cl^-輸送ATPaseに関する分子生物学的研究" (財)薬学研究奨励財団「薬学研究の進歩・研究成果報告集」. 7. 137-148 (1991)

    • Related Report
      1992 Annual Research Report
  • [Publications] Ikeda,M.,Satoh,S.,Maeshima,M.,Mukohata,Y.,&Moritani,C.: "A vacuolar ATPase and pyrophosphatase in Acetabularia acetabulum" Biochim. Biophys. Acta. 1070. 77-82 (1991)

    • Related Report
      1992 Annual Research Report
  • [Publications] Ohhashi,T.,Katsu,T.&Ikeda,M.: "Improvement of reconstitution of the Cl^--translocating ATPase isolated from Acetabularia acetabulum into liposomes and several anion pump characteristics" Biochim. Biophys. Acta. 1106. 165-170 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] 池田 己喜子,大森 晋爾: "生体膜の存在するアニオン輸送系の研究-Cl^-輸送性ATPase及び硫酸イオン輸送系の解析" 日本学術振興会「学術月報」,. (1993)

    • Related Report
      1992 Annual Research Report
  • [Publications] Moritani,C.,Yamauchi,M.,de Groot,E.J.,Oesterhelt,D. & Ikeda,M.: "Acetabularia is equipped with sulfate permease:amplification of a 339 bp and a 552 bp fragment encoding a putative cysA and a sulfate binding protein" Planta,. (1993)

    • Related Report
      1992 Annual Research Report
  • [Publications] Moritani,C.,Ohhayashi,T.,Lottspeich,F.,Oesterhelt,D. & Ikeda,M.: "Cloning and sequencing of the gene encording the subunit of the Cl^--trnaslocating ATPase of Acetabularia acetabulum" Biochim.Biophys.Acta,. (1993)

    • Related Report
      1992 Annual Research Report

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Published: 1992-04-01   Modified: 2016-04-21  

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