1990 Fiscal Year Final Research Report Summary
Oligosaccharide Structure and Biological Activity of Erythropoietin
| Project/Area Number |
63480504
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
代謝生物化学
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| Research Institution | Nagoya City University College of Nursing |
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Principal Investigator |
TAKAHASHI Noriko Nagoya City Univ. College of Nursing Prof., 看護学科, 教授 (90079989)
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| Co-Investigator(Kenkyū-buntansha) |
ISHIHARA Hideko Nagoya City Univ. Faculty Pharm. Associate Prof., 薬学部, 助教授 (90080192)
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| Project Period (FY) |
1988 – 1989
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| Keywords | Erythropoietin / recombinant EPO / BHK cell / Asn-linked oligosaccnarides |
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| Research Abstract |
The structures of the N-linked oligosaccharides of the urinary erythropoietin (u-EPO) purified from urine of aplastic anemic patients were analyzed and compared with those for recombinant erythropoietin (r-EPO) prepared with Baby Hamster Kidney (BHK) cells. Asparagine-linked neutral oligosaccharides were related from each EPO protein by N-oligosaccharide glycopeptidase (almond) digestion. The reducing ends of the oligosaccharide chains thus obtained were aminated with a fluorescent reagent, 2-aminopyridine, and the mixture of pyridylamino derivatives of the oligosaccharides was separated by High-Performance Liquid Chromatography (HPLC) on an ODS silica column. More than 8 and 13 kinds of oligosaccharide fractions for u-EPO and r-EPO (BHK), respectively, were completely separated by the one-step HPLC procedure. The structure of each oligosaccharide thus isolated was analyzed by a combination of sequential exoglycosidase digestion and another kind of HPLC with an amide-silica column. Furthermore, ^IH NMR spectroscopy and methylation analyses were carried out in the case of r-EPO (BHK). It has been found that (1) significantly different distributions of oligosaccharide components exist in u-EPO preparations isolated from different individuals, whereas the distribution of oligosaccharide components is quite similar for different r-EPO (BHK) preparations, (2) in r-EPO (BHK) a tetraantennary oligosaccharide with a fucose residue is predominant and various higher molecular weight tetraantennary oligosaccharides with one to three Gal( 1-4) GlcNAc repeating units exist, (3) all of the eight kinds of oligosaccharides of u-EPO also exist in r-EPO (BHK), although the distribution of the oligosaccharides is different for the two proteins, and (4) oligosaccharides with unusualstructures containing the Gal (1-3) GlcNAc group exist in both u- and r-EPO (BHK).
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Research Products
(16 results)
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[Publications] Goto, M., Akai, K., Murakami, A., Hashimoto, C., Tsuda, E., Ueda, M Kawanishi, G., Takahashi, N., Ishimoto, A., Chiba, H., & Sakaki, R.: "Production of recombinant human erythropoietin in mammalian cells : Host-cell dependency of the biological activity of the cloned glycoprotein." Biotechnology. 6. 67-71 (1988)
Description
「研究成果報告書概要(欧文)」より
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[Publications] Tomiya, N., Awaya, J., Kurono, M., Endo, S., Arata, Y., & Takahashi, N.: "Analyses of N-linked oligosaccharides using a two dimensional mapping technique." Anal. Biochem. 171. 73-90 (1988)
Description
「研究成果報告書概要(欧文)」より
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[Publications] Tsuda, E., Goto, M., Murakami, A., Akai, K., Ueda, M., Kawanishi, G., Takahashi, N., Sasaki, R., Chiba, H., Ishihara, H., Mori, M., Tejima, S., Endo, S., & Arata, Y.: "Comparative structural study of N-linked oligosaccharides of urinary and recombinant erythropoietins." Biochemistry. 27. 5646-5654 (1988)
Description
「研究成果報告書概要(欧文)」より
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[Publications] Tomiya, N., Yamaguchi, T., Awaya, J., Kurono, M., Endo, S., Arata, Y., Takahashi, N., Ishihara, H., Mori, M., & Tejima, S.: "Structural analyses of asparagine-linked oligosaccharides of porcine pancreatic kallikrein." Biochemistry. 27. 7146-7154 (1988)
Description
「研究成果報告書概要(欧文)」より
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[Publications] Yonekura, H., Oguri, K., Itoh, H., Tanabe, K., Takahashi, N., Nakanishi, Y., & Okayama, M.: "Sulfated glycoproteins synthesized by the corneal epithelium of chick embryo having the same molecular weights as cytokeratin polypeptides." J. Biochem. 104. 648-657 (1988)
Description
「研究成果報告書概要(欧文)」より
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[Publications] Gu, J., Matsuda, T., Nakamura, R., Ishiguro, H., Okubo, I., Sasaki, M., & Takahashi, N.: "Chemical deglycosylation of hen ovomucoid : Protective effect of carbohydrate moiety on tryptic hydrolysis and heat denaturation." J. Biochem.106. 66-70 (1989)
Description
「研究成果報告書概要(欧文)」より
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[Publications] Yamazumi, K., Shimura, K., Terukina, S., Takahashi, N., & Matsuda, M.: "A gamma-methionine-310 to threonine substitution and consequent Nglycosylation at gamma-asparagine-308 identified in a congenital dysfibrinogenemia associated with posttraumatic bleeding, Fibrinogen Asahi." J. Clin. Invest. 83. 1590-1597 (1989)
Description
「研究成果報告書概要(欧文)」より
