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Enzymatic properties of a phospholipase A_2 from vascular smooth muscles and its physiological significance

Research Project

Project/Area Number 04680190
Research Category

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

Allocation TypeSingle-year Grants
Research Field 代謝生物化学
Research InstitutionOsaka University

Principal Investigator

TOJO Hiromasa  Osaka University Medical School, Department of Molecular Physiological Chemistry, Lecturer, 大阪大学医学部, 講師 (90135707)

Project Period (FY) 1992 – 1993
Project Status Completed (Fiscal Year 1993)
Budget Amount *help
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1993: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1992: ¥1,300,000 (Direct Cost: ¥1,300,000)
Keywordsphospholipase A_2 / bovine aorta / phospholipase B / phospholipids / 血管平滑筋 / 牛大動脈中膜
Research Abstract

1. Freshly prepared extracts of bovine aorta contained a minimal phospholipase A (PLA) activity towards either various combinations of different phospholipids and detergents or phospholipid dispersions. However, incubation of the extracts at 30゚C under sterile conditions led to a gradual increase in Ca^<2+>-dependent PLA activity towards phosphatidylethanolamine (PE) dispersions after a 6-hour lag, and it reached a maximum after 48h. This activation of PLA activity absolutely required the presence of Ca^<2+> ions. Other divalent cations, Sr^<2+> and Mg^<2+> also were effective in this order, but to a less extent than Ca^<2+> ions. Guanine nucleotides, GTP, GMP, cGMP, and guanosine, enhanced the activation rate, whereas ATP and cAMP suppressed it. Interestingly, NO-forming reagents, NaN_3, NaNO_2, and isosorbide dinitrate, inhibited the activation. Serine protease inhibitors markedly inhibited the activation, suggesting the activation mechanism through the limited proteolysis of an inactive prophospholipase A.
2. We purified to near homogeneity bovine aorta PLA, which had been previously activated at 4゚C, through the sequential use of Q-Sepharose, phenyl-Sepharose, Sepharose 4B, and Cosmosil 5C_8-300 chromatographies. The purified enzyme was characterized as follows : a, The enzyme prefer PE dispersions over other classes of phospholipid. Anionic detergent inhibited the enzyme activity. : b, The molecular mass of the enzyme was found to be about 30 kDa. c, Calcium ions were essential for activity. d, Fatty acids were released from both sn-1 and sn-2 positions of mixed acyl phospholipids by the enzyme action. e, Bovine aorta PLA was immunochemically distinct from group I, and II phospholipases A_2.

Report

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

    (28 results)

All Other

All Publications (28 results)

  • [Publications] Tojo,H.: "Analytical and micropreparative high performance gel chromatography of proteins with a short column" J.Chromatogr.605. 205-213 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Shinohara,H.et al.: "Group II phospholipase A_2 induced by interleukin 1 in cultured rat gingival fibroblasts" FEBS Lett.304. 69-72 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Tojo,H.et al.: "High performance liquid chromatographic assay of phospholipases" J.Lipid Res.34. 837-844 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Tojo,H.et al: "Purification and characterization of guinea pig gastric phospholipase A_2 of the pancreatic type" Eur.J.Biochem.215. 81-90 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Zhao,Y.et al: "Cloning and expression of phospholipase A_2 from guinea pig gastric mucosa" Eur.J.Biochem.215. 91-97 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Minami,T.et al.: "Purification and characterization of a phospholipase A_2 from human ileal mucosa" Bichim.Biophys.Acta. 1170. 125-130 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Shinohara, H., Amabe, Y., Komatsubara, T., Tojo, H., Okamoto, M., Wakano, Y., and Ishida, H.: "Group II phospholipase A_2 induced by interleukin 1 in cultured rat ginagival fibroblasts." FEBS Lett.304. 69-72 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Tojo, H., Horiike, K., Ishida, T., Kobayashi, T., Nozaki, M., and Okamoto, M.: "Analytical and micropreparative high-performance gel chromatography of proteins with a short column." J.Chromatogr.605. 205-213 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Minami, T., Tojo, H., Shinomura, Y., Tarui, S., and Okamoto, M.: "Raised serum activity of phospholipase A_2 immunochemically related to group II enzyme in inflammatory bowel disease." Gut. 33. 914-921 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Tojo, H., Ono, T., and Okamoto, M.: "High performance liquid chromatographic assay of phospholipases. Application to spectrophotometric detection of rat phospholipase A_2 isozymes." J.Lipid Res.34. 837-844 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Tojo, H., Zhao, Y., and Okamoto, M.: "Purification characterization of guinea pig gastric phospholipase A_2 of the pancreatic type." Eur.J.Biochem.215. 81-90 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Zhao, Y., Tojo, H., Nonaka, Y., and Okamoto, M.: "Cloning and expression of phospholipase A_2 from guinea pig gastric mucosa, its induction and secretion in vivo by carbachol." Eur.J.Biochem.215. 91-97 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Minami, T., Tojo, H., Shinomura, Y., Komatsubara, T., Matsuzawa, Y., and Okamoto, M.: "Elevation of group II phospholipase A_2 protein in sera of patients with Crohn's disease and ulcerative colitis." Am.J.Gastroenterol. 88. 1076-1080 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Minami, T., Tojo, H., Shinomura, Y., Matsuzawa, Y., and Okamoto, M.: "Purification and characterization of a phospholipase A_2 from human ileal mucosa." Biochem.Biophys.Acta. 1170. 125-130 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Ishida, H., Shinohara, H., Amabe, Y., Tojo, H., Nagata, T., and Wakano, Y.: "Effects of interleukin 1beta, tumor necrosis factor and transforming growth factor on group II phospholipase A_2 activity in rat gingival fibroblasts." J.Periodont.Res.28. 517-520 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Yamamoto, K., Shinomura, Y., Tojo, H., Okamoto, M., and Tarui, S.: "Serum pancreatic phospholipase A_2 in acute pancreatitis and after endoscopic retrograde pancreatography." Gastroenterol.Jpn.28. 679-686 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Tojo,H et al.: "High performance liquid chromatographic assay of phospholipases.Application to spectrophotometric detection of PLA_2" J.Lipid Res.34. 837-844 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] Tojo,H et al.: "Purification and characterization of guinea pig gastric phospholipase A_2 of the pancreatic type" Eur.J.Biochem.215. 81-90 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] Zhao,Y.et al.: "Cloning and expression of phospholipase A_2 from guinea pig gastric mucosa,its induction in vivo and secretion by carbachol" Eur.J.Biochem.215. 91-97 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] Minami,T.et al.: "Purification of characterization of a phospholipase A_2 from human ileal mucosa" Biochim.Biophys.Acta. 1170. 125-130 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] Minami,T.et al.: "Elevation of group II phospholipase A_2 protein in sera of patients with Crohn's disease and ulcerative colitis" Am.J.Gastroenterol.88. 1076-1080 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] Yamamoto,K.et al.: "Serum pancreatic phospholipase A_2 in acute pancreatitis and after endoscopic retrograde pancreatography" Gastroenterol.Jpn.28. 679-686 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] Tojo,H.,et al.: "Analytical and micropreparative high-performance gel chromatography of proteins with a short column" J.Chromatogr.605. 205-213 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] Minami.T.,et al.: "Raised serum activity of phospholipase A_2 immunochemically related to group II enzyme in inflammatory bowel disease" GUT. 33. 914-921 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] Shinohara,H.,et al.: "Group II phospholipase A_2 induced by interleukin 1 in cultured rat gingival fibroblasts." FEBS LETT.304. 69-72 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] Tojo,H.,et al.: "High performance liquid chromatographic assay of phospholipases.application to spectrophotometric detection of rat phospholipase A_2 isozymes" J.Lipid Res.(1993)

    • Related Report
      1992 Annual Research Report
  • [Publications] Minami,T.,et al.: "Elevation of phospholipase A_2 protein in sera of patients with Crohns disease and ulcerative colitis" Am.J.Gastroenterol.

    • Related Report
      1992 Annual Research Report
  • [Publications] Yamamoto,K.,et al.: "Serum pancreatic phospholipase A_2 and prophospholipase A_2 in acute pancreatitis and after endoscopic retrograde paccreatography" Gasroenterologia Japonica.

    • Related Report
      1992 Annual Research Report

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

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