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Chemo-enzymatic synthesis and structure-activity study of artificially N-glycosylated eel calcitonin derivatives with a complex type oligosaccharide

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Abstract

Starting from N-glycosylated eel calcitonin derivatives that contain an N-acetyl-D-glucosamine residue specifically at the 3rd, 14th, 20th or 26th amino acid residue, corresponding glycopeptides with a complex-type oligosaccharide attached to the respective amino acid residue were synthesized by means of a transglycosylation reaction catalyzed by an endo-β-N-acetylglucosaminidase from Mucor hiemalis. The use of a recombinant enzyme and an excess of a glycosyl donor led to a yield in excess of 60%. Calcitonin derivatives containing truncated oligosaccharides were also prepared via digestion of the complex-type N-glycan with exoglycosidases. Using these N-glycosylated calcitonin derivatives, the effect of carbohydrate structure and glycosylation site on the three-dimensional structure and the biological activity of the peptide were studied. The conformation of the peptide backbone did not change irrespective of the carbohydrate structure or the glycosylation site. However, hypocalcemic activity, calcitonin-receptor binding activity and the biodistribution of the derivatives were affected by the glycosylation and were dependent on both the carbohydrate structure and the glycosylation site. Although the larger oligosaccharides tended to hinder receptor binding, the biodistribution altered by N-glycosylation appeared to enhance the hypocalcemic activity in some cases, and the magnitude of the effect was dependent on the site of glycosylation. Published in 2003.

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Authors and Affiliations

  1. The Noguchi Institute, 1-8-1, Kaga, Itabashi, Tokyo, 173-0003, Japan

    Katsuji Haneda, Eiichi Yoshino, Midori Takeuchi, Toshiyuki Inazu & Kazunori Toma

  2. Asahi Kasei Co., Fuji, Shizuoka, 416-8501, Japan

    Mizuka Tagashira & Hideki Iijima

  3. Department of Applied Chemistry, School of Engineering, Tokai Universtiy, Hiratsuka, Kanagawa, 259-1292, Japan

    Toshiyuki Inazu

  4. First Laboratory for Pharmacology, Asahi Kasei Co., Tagata, Shizuoka, 410-2321, Japan

    Yukihiro Isogai & Masayuki Hori

  5. High Energy Medical Research Center, Fukui Medical University, Yoshida, Fukui, 910-1193, Japan

    Shinji Takamatsu & Yasuhisa Fujibayashi

  6. Central Laboratories for Key Technology, Kirin Brewery Co., Kanazawa, Yokohama, Kanagawa, 236-0004, Japan

    Kazuo Kobayashi & Makoto Takeuchi

  7. Graduate School of Biostudies, Kyoto University, Sakyo, Kyoto, 606-8502, Japan

    Kenji Yamamoto

Authors
  1. Katsuji Haneda
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  2. Mizuka Tagashira
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  3. Eiichi Yoshino
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  4. Midori Takeuchi
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  6. Kazunori Toma
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  7. Hideki Iijima
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  8. Yukihiro Isogai
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  9. Masayuki Hori
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  11. Yasuhisa Fujibayashi
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  12. Kazuo Kobayashi
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Correspondence to Katsuji Haneda.

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Haneda, K., Tagashira, M., Yoshino, E. et al. Chemo-enzymatic synthesis and structure-activity study of artificially N-glycosylated eel calcitonin derivatives with a complex type oligosaccharide. Glycoconj J 21, 377–386 (2004). https://doi.org/10.1023/B:GLYC.0000046277.92806.74

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  • DOI: https://doi.org/10.1023/B:GLYC.0000046277.92806.74

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