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Sphingolipid Engineering Using Marine Resources

Research Project

Project/Area Number 13460044
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field 応用微生物学・応用生物化学
Research InstitutionKyushu University

Principal Investigator

ITO Makoto  Kyushu University, Faculty of Agriculture, Department of Bioscience and Biotechnology, Professor, 大学院・農学研究院, 教授 (40253512)

Project Period (FY) 2001 – 2002
Project Status Completed (Fiscal Year 2002)
Budget Amount *help
¥13,400,000 (Direct Cost: ¥13,400,000)
Fiscal Year 2002: ¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 2001: ¥10,400,000 (Direct Cost: ¥10,400,000)
Keywordssphingolipids / sphingolipid ceramide N-deacylase / endoglycoceramidase / marine bacteria / marine invertebrate / gene cloning / ceramidase / sphingomyelinase / 海洋細菌 / スフィンゴ糖脂質 / レクチン / 代謝酵素遺伝子 / 糖鎖工学 / 縮合反応(加水分解逆反応) / 糖転移反応 / 海洋生物資源 / 代謝酵素
Research Abstract

We report here the purification, characterization, and cDNA cloning of a novel N- acetylgalactosamine-specific lectin from starfish, Asterina pectinifera. The purified lectin showed 19-kDa, 41-kDa and 60-kDa protein bands on SDS-PAGE, possibly corresponding to a monomer, homodimer and homotrimer. The hemagglutination activity of the lectin was specifically inhibited by N-acetylgalactosamine, Tn antigen and blood group A trisaccharide, but not by N-acetylglucosamine, galactose, galactosamine or blood group B trisaccharide. cDNA cloning of the lectin showed that the protein is composed of 168 amino acids including a signal sequence of 18 residues, and possesses the typical C-type lectin motif. These findings indicate that the protein is a C-type lectin.
We report here the purification, characterization, molecular cloning, and expression of the gene encoding the enzyme from marine bacterium, Sewanella alga G8. The enzyme was purified by 1,600fold with an overall yield of 24% from a culture supernatant of strain G8. After being stained with a silver staining solution, molecular mass of the purified enzyme was estimated to be 75 kDa on SDS-polyacrylamide gel electrophoresis. The open reading frame of 2,976 nucleotides encoded a polypeptide of 992 amino acids including a signal sequence of 35 residues, 59 residues of which matched the amino acid sequence determined for the purified enzyme. The molecular weight of the mature enzyme was estimated to be 105,994 from the deduced amino acid sequence.

Report

(3 results)
  • 2002 Annual Research Report   Final Research Report Summary
  • 2001 Annual Research Report
  • Research Products

    (20 results)

All Other

All Publications (20 results)

  • [Publications] Y.Horibata 他: "Transglycosylation of reverse hydrolysis reaction of endoglycoceramidase from the Jellyfish, Cyanea nozakii"Journal of Biochemistry. 130(2). 263-268 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] T.Nkagawa, M.Ito: "Distribution of neutral/alkaline ceramidase in marine invertebrates revealed by fluorescence-labeled ceramide"Fisheries Science. 67. 544-546 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] N.Sueyoshi: "Molecular cloning and expression of Mn2+ dependent sphingomyelinase/hemolysin of an aquatic bacterium Pseudomonas sp."Journal of Bacteriology. 184(2). 540-546 (2002)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] M.Kakiuchi 他: "Purification, characterization, and cDNA cloning of α-N-acetylgalactosaminidase-specific lectin from starfish, Asterrina pectinifera"Glycobiology. 12(2). 85-94 (2002)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] M.Furusato 他: "Molecular cloning and characterization of sphingolipid ceramide N-deacylase from a marine bacterium, Shewanella alga G8"Journal of Biological Chemistry. 277(19). 17300-17307 (2002)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] T.Sumida 他: "Utilization of ganglioside-degrading Paenibacillus sp. strain TS12 for production of glucosylceramide"Applied and Environmental Microbiology. 68(11). 5241-5248 (2002)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Yasuhiro Horibata, Hideyoshi Higashi, and Makoto Ito: "Transglaycosylation and Reverse Hydrolysis Reactions of Endoglycoceramidase from the Jellyfish, Cyanea nozakii"Journal of Biochemistry. 130(2). 263-268 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Tetsuto Nakagawa and Makoto Ito: "Distribution of Neutral/Alkaline Ceramidase in Marine Invertebrates Revealed by Fluorescence-labeled Ceramide"Fisheries Science. 67. 544-546 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Noriyuki Sueyoshi, Katsuhiro Kita, Nozomu Okino, Keishi Sakaguchi, Takashi Nakamura and Makoto Ito: "Molecular Cloning and Expression of Mn^<2+>- Dependent Sphingomyelinase/Hemolysin of an Aquatic Bacterium, Pseudomonas sp. Strain TK4"Journal of Bacteriology. 184(2). 540-546 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Mari Kakiuchi, Nozomu Okino, Noriyuki Sueyoshi, Sachiyo Ichinose, Akira Omori, Shuichiro Kawabata, Kuniko Yamaguchi and Makoto Ito: "Purification, Characterization, and cDNA cloning of a-N-acetylgalactosamine-specific lectin from starfish, Asterina pectinifera"Glycobiology. 12(2). 85-94 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Masako Furusato, Noriyuki Sueyoshi, Susumu Mitstutake, Keishi Sakaguchi, Katsuhiro Kita, Nozomu Okino, Sachiyo Ichinose, Akira Omori, and Makoto Ito: "Molecular Cloning and Characterization of Sphingolipid Ceramide N- Deacylase from a Marine Bacterium, Shewanella alga G8"Journal of Biological Chemistry. 277(19). 17300-17307 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Tomomi Sumida, Noriyuki Sueyoshi, and Makoto Ito: "Utilization of Ganglioside-Degrading Paenibacillus sp. Strain TS12 for Production of Glucosylceramide"Applied and Environmental Microbiology. 68(11). 5241-5248 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Furusato, M. et al.: "Molecular cloning and characterization of sphingolipid ceramide N-deacylase from a marine bacterium, Shewanella alga G8"Journal of Biological Chemistry. 277(19). 17300-17307 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Kakiuchi, M. et al.: "Purification, characterization, and cDNA cloning of α-N-acetylgalactosamine-specific lectin from starfish, Asterina pectinifera"Glycobiology. 12(2). 85-94 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Horibata, Y. et al.: "Transglycosylation and reverse hydrolysis reactions of endoglycoceramidase from the Jellyfish, Cyanea nozakii"Journal of Biochemistry. 130(2). 263-268 (2001)

    • Related Report
      2002 Annual Research Report
  • [Publications] Sueyoshi, N. et al.: "Molecular cloning and expression of Mn2+-dependent sphingomyelinase/hemolysin of an aquatic bacterium, Pseudomonas sp."Journal of Bacteriology. 184(2). 540-546 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Horibata, Y. et al.: "Transglycosylation and reverse hydrolysis reactions of endoglycoceramidse from the jellyfish, Cyanea nozaki"Journal of Biochemistry. 130. 263-268 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Nakagawa et al.: "Distribution of neutral/alkaline ceramidases in marine invertebrates revealed by fluorescence-labeled ceramide"Fisheries Science. 67. 544-546 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Sueyoshi, N. et al.: "Molecular cloning and expression of Mn2+-dependent sphingomyelinase/hemolysin of an aquatic bacterium Pseudomonas sp. Strain TK4"Journal of Bacteriology. 184. 540-546 (2002)

    • Related Report
      2001 Annual Research Report
  • [Publications] Furusato, M.: "Molecular cloning and characterization of sphingolipid ceramide N-deacylase"Journal of Biological Chemistry. 277(in press). (2002)

    • Related Report
      2001 Annual Research Report

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Published: 2001-04-01   Modified: 2016-04-21  

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