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Study on suppression of oxidative damage by copper-binding protein

Research Project

Project/Area Number 08457055
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field Pathological medical chemistry
Research InstitutionWakayama Medical College

Principal Investigator

NISHIKIMI Morimitsu  Wakayama Medical College, Medicine, Professor, 医学部, 教授 (20022816)

Co-Investigator(Kenkyū-buntansha) MATSUI LEE In Sook  Wakayama Medical College, Medicine, Research associate, 医学部, 助手 (50209475)
SHIRAISHI Noriyuki  Wakayama Medical College, Medicine, Lecturer, 医学部, 講師 (30133169)
Project Period (FY) 1996 – 1997
Project Status Completed (Fiscal Year 1997)
Budget Amount *help
¥4,700,000 (Direct Cost: ¥4,700,000)
Fiscal Year 1997: ¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 1996: ¥2,400,000 (Direct Cost: ¥2,400,000)
KeywordsCopper-binding protein / S100b protein / Hemolysis / Reactive oxygen species / Antioxidant / Escherichia coli / 銅結合性タンパク質 / キレート / アスコルビン酸
Research Abstract

This study was aimed at elucidation of the importance of copper-binding protein in suppression of copper-induced cellular oxidative damage. We have isolated from bovine brain a protein with a high capacity to inhibit the copper ion-catalyzed oxidation of L-ascorbate and identified it as S100b protein, an EF-hand calcuim binding protein, by sequencing its proteolytis peptides. Copper binding studies showed that this protein has four copper binding sites per dimeric protein molecule with a dissociation constant of 0.46 muM and that in the presence of L-ascorbate, copper ions bind to a total of six binding sites with a great increase in affinity. Furthermore, we examined whether S100b protein can prevent copper-induced cell damage. Bovine S100b protein was found to suppress dose-dependently the hemolysis of mouse erythrocytes indused by CuCl_2. We transformed Escherichia coli cells with pGEX-5X-3 vector containing a cDNA for rat S100b protein, so that this protein could be extressed as a fusion protein with glutathione S-transferase.The transformed cells were demonstrated to be markedly resistant to a treatment with CuCl_2 plus H_2O_2 as compared with the control cells expressing glutathions S-transferase alone. A study by gel chromatography and affinity chromatography for S100beta fusion protein showed that the polymeric form of the fusion protein expressed in the cells mainly bound the copper that had been incorporated into the cells. These results indicate S100b protein does suppress oxidative cell damage by sequestering copper ions.

Report

(3 results)
  • 1997 Annual Research Report   Final Research Report Summary
  • 1996 Annual Research Report
  • Research Products

    (6 results)

All Other

All Publications (6 results)

  • [Publications] T.Nishikawa: "Identification of S100b protein as copper-binding protein and its suppression of copper-induced cell damage" Journal of Biological Chemistry. 272・37. 23037-23041 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] 錦見 盛光: "銅で誘起される酸化障害の防御系" フリーラジカルの臨床. 12. 9-14 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] T.Nishikawa, I.S.Matsui Lee, N.Shiraishi, T.Ishikawa, Y.Ohta, and M.Nishikimi: "Indentification of S100b protein as copper-binding protein and its suppression of copper-induced cell damage" J.Biol.Chem.272(3). 23037-23041 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] M.Nishikimi: "Defense system against copper-induced oxdative damage" Free Radicals in Clinical Medicine (in Japanese). 12. 9-4 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] T.Nishikawa: "Identification of S100b protein as copper-binding protein and its suppression of copper-induced cell damage" Journal of Biological Chemistry. 272・37. 23037-23041 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] 錦見盛光: "銅で誘起される酸化障害の防御系" フリーラジカルの臨床. 12. 9-4 (1997)

    • Related Report
      1997 Annual Research Report

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Published: 1996-04-01   Modified: 2016-04-21  

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