Glycoconjugate Journal

, Volume 27, Issue 1, pp 125–132 | Cite as

Syntheses of mucin-type O-glycopeptides and oligosaccharides using transglycosylation and reverse-hydrolysis activities of Bifidobacterium endo-α-N-acetylgalactosaminidase

  • Hisashi Ashida
  • Hayato Ozawa
  • Kiyotaka Fujita
  • Shun’ichi Suzuki
  • Kenji Yamamoto
Article
First Online:
Received:
Revised:
Accepted:

Abstract

Endo-α-N-acetylgalactosaminidase catalyzes the release of Galβ1-3GalNAc from the core 1-type O-glycan (Galβ1-3GalNAcα1-Ser/Thr) of mucin glycoproteins and synthetic p-nitrophenyl (pNP) α-linked substrates. Here, we report the enzymatic syntheses of core 1 disaccharide-containing glycopeptides using the transglycosylation activity of endo-α-N-acetylgalactosaminidase (EngBF) from Bifidobacterium longum. The enzyme directly transferred Galβ1-3GalNAc to serine or threonine residues of bioactive peptides such as PAMP-12, bradykinin, peptide-T and MUC1a when Galβ1-3GalNAcα1-pNP was used as a donor substrate. The enzyme was also found to catalyze the reverse-hydrolysis reaction. EngBF synthesized the core 1 disaccharide-containing oligosaccharides when the enzyme was incubated with either glucose or lactose and Galβ1-3GalNAc prepared from porcine gastric mucin using bifidobacterial cells expressing endo-α-N-acetylgalactosaminidase. Synthesized oligosaccharides are promising prebiotics for bifidobacteria.

Keywords

Bifidobacteria Endoglycosidase GH101 Mucin O-glycan Prebiotics 
This is a preview of subscription content, log in to check access.

Notes

Acknowledgments

We thank Dr. H. Ishida and Dr. M. Kiso (Gifu University), Dr. M. Yamaguchi (Wakayama University) and Dr. T. Yamanoi (the Noguchi Institute) for MS analyses. This work was supported in part by grant-in-aids from the Promotion of Basic Research Activities for Innovative Biosciences (PROBRAIN) and from Ajinomoto Co., Inc.

References

  1. 1.
    Rich, J.R., Withers, S.G.: Emerging methods for the production of homogeneous human glycoproteins. Nat. Chem. Biol. 5, 206–215 (2009). doi: 10.1038/nchembio.148 CrossRefPubMedGoogle Scholar
  2. 2.
    Ashida, H., Tsuji, Y., Yamamoto, K., Kumagai, H., Tochikura, T.: Transglycosylation activity of endoglycoceramidase from Corynebacterium sp. Arch. Biochem. Biophys. 305, 559–562 (1993). doi: 10.1006/abbi.1993.1461 CrossRefPubMedGoogle Scholar
  3. 3.
    Yamamoto, K., Kadowaki, S., Watanabe, J., Kumagai, H.: Transglycosylation activity of Mucor hiemalis endo-β-N-acetyl-glucosaminidase which transfers complex oligosaccharides to the N-acetylglucosamine moieties of peptides. Biochem. Biophys. Res. Commun. 203, 244–252 (1994). doi: 10.1006/bbrc.1994.2174 CrossRefPubMedGoogle Scholar
  4. 4.
    Ashida, H., Yamamoto, K., Murata, T., Usui, T., Kumagai, H.: Characterization of endo-α-N-acetylgalactosaminidase from Bacillus sp. and syntheses of neo-oligosaccharides using its transglycosylation activity. Arch. Biochem. Biophys. 373, 394–400 (2000). doi: 10.1006/abbi.1999.1565 CrossRefPubMedGoogle Scholar
  5. 5.
    Ashida, H., Yamamoto, K., Kumagai, H.: Enzymatic syntheses of T antigen-containing glycolipid mimicry using the transglycosylation activity of endo-α-N-acetylgalactosaminidase. Carbohydr. Res. 330, 487–493 (2001). doi: 10.1016/S0008-6215(01)00008-8 CrossRefPubMedGoogle Scholar
  6. 6.
    Umekawa, M., Huang, W., Li, B., Fujita, K., Ashida, H., Wang, L.X., et al.: Mutants of Mucor hiemalis endo-β-N-acetylglucosaminidase show enhanced transglycosylation and glycosynthase-like activities. J. Biol. Chem. 283, 4469–4479 (2008). doi: 10.1074/jbc.M707137200 CrossRefPubMedGoogle Scholar
  7. 7.
    Umemura, M., Itoh, M., Makimura, Y., Yamazaki, K., Umekawa, M., Masui, A., et al.: Design of a sialylglycopolymer with a chitosan backbone having efficient inhibitory activity against influenza virus infection. J. Med. Chem. 51, 4496–4503 (2008). doi: 10.1021/jm8000967 CrossRefPubMedGoogle Scholar
  8. 8.
    Fujita, K., Oura, F., Nagamine, N., Katayama, T., Hiratake, J., Sakata, K., et al.: Identification and molecular cloning of a novel glycoside hydrolase family of core 1 type O-glycan-specific endo-α-N-acetylgalactosaminidase from Bifidobacterium longum. J. Biol. Chem. 280, 37415–37422 (2005). doi: 10.1074/jbc.M506874200 CrossRefPubMedGoogle Scholar
  9. 9.
    Anderson, K., Li, S.C., Li, Y.T.: Diphenylamine-aniline-phosphoric acid reagent, a versatile spray reagent for revealing glycoconjugates on thin-layer chromatography plates. Anal. Biochem. 287, 337–339 (2000). doi: 10.1006/abio.2000.4829 CrossRefPubMedGoogle Scholar
  10. 10.
    Kuwasako, K., Kitamura, K., Ishiyama, Y., Washimine, H., Kato, J., Kangawa, K., et al.: Purification and characterization of PAMP-12 (PAMP[9–20]) in porcine adrenal medulla as a major endogenous biologically active peptide. FEBS Lett. 414, 105–110 (1997). doi: 10.1016/S0014-5793(97)00971-X CrossRefPubMedGoogle Scholar
  11. 11.
    Elliott, D.F., Horton, E.W., Lewis, G.P.: The isolation of bradykinin, a plasma kinin from ox blood. Biochem. J. 78, 60–65 (1961)PubMedGoogle Scholar
  12. 12.
    Gendler, S.J., Lancaster, C.A., Taylor-Papadimitriou, J., Duhig, T., Peat, N., Burchell, J., et al.: Molecular cloning and expression of human tumor-associated polymorphic epithelial mucin. J. Biol. Chem. 265, 15286–15293 (1990)PubMedGoogle Scholar
  13. 13.
    Pert, C.B., Hill, J.M., Ruff, M.R., Berman, R.M., Robey, W.G., Arthur, L.O., et al.: Octapeptides deduced from the neuropeptide receptor-like pattern of antigen T4 in brain potently inhibit human immunodeficiency virus receptor binding and T-cell infectivity. Proc. Natl Acad. Sci. USA 83, 9254–9258 (1986). doi: 10.1073/pnas.83.23.9254 CrossRefPubMedGoogle Scholar
  14. 14.
    Ashida, H., Maki, R., Ozawa, H., Tani, Y., Kiyohara, M., Fujita, M., et al.: Characterization of two different endo-α-N-acetylgalactosaminidases from probiotic and pathogenic enterobacteria. Bifidobacterium longum and Clostridium perfringens. Glycobiology 18, 727–734 (2008). doi: 10.1093/glycob/cwn053 Google Scholar
  15. 15.
    Amano, K., Chiba, Y., Kasahara, Y., Kato, Y., Kaneko, M.K., Kuno, A., et al.: Engineering of mucin-type human glycoproteins in yeast cells. Proc. Natl Acad. Sci. USA 105, 3232–3237 (2008). doi: 10.1073/pnas.0710412105 CrossRefPubMedGoogle Scholar
  16. 16.
    Ten Hagen, K.G., Fritz, T.A., Tabak, L.A.: All in the family: the UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferases. Glycobiology 13, 1R–16R (2003). doi: 10.1093/glycob/cwg007 CrossRefPubMedGoogle Scholar
  17. 17.
    Ajisaka, K., Miyasato, M., Ishii-Karakasa, I.: Efficient synthesis of O-linked glycopeptide by a transglycosylation using endo α-N-acetylgalactosaminidase from Streptomyces sp. Biosci. Biotechnol. Biochem. 65, 1240–1243 (2001). doi: 10.1271/bbb.65.1240 CrossRefPubMedGoogle Scholar
  18. 18.
    Tarp, M.A., Clausen, H.: Mucin-type O-glycosylation and its potential use in drug and vaccine development. Biochim. Biophys. Acta 1780, 546–563 (2008)PubMedGoogle Scholar
  19. 19.
    Imamura, A., Ando, H., Ishida, H., Kiso, M.: Di-tert-butylsilylene-directed α-selective synthesis of 4-methylumbelliferyl T-antigen. Org. Lett. 7, 4415–4418 (2005). doi: 10.1021/ol051592z CrossRefPubMedGoogle Scholar
  20. 20.
    Imamura, A., Kimura, A., Ando, H., Ishida, H., Kiso, M.: Extended applications of di-tert-butylsilylene-directed α-predominant galactosylation compatible with C2-participating groups toward the assembly of various glycosides. Chemistry (Easton) 12, 8862–8870 (2006)Google Scholar
  21. 21.
    Sato, T., Imamura, A., Ando, H., Ishida, H., Kiso, M.: Di-tert-butylsilylene-directed α-selective synthesis of p-nitrophenyl T-antigen analogues. Glycoconj. J. 26, 83–98 (2009). doi: 10.1007/s10719-008-9168-y CrossRefPubMedGoogle Scholar
  22. 22.
    Wada, J., Suzuki, R., Fushinobu, S., Kitaoka, M., Wakagi, T., Shoun, H., et al.: Purification, crystallization and preliminary X-ray analysis of the galacto-N-biose-/lacto-N-biose I-binding protein (GL-BP) of the ABC transporter from Bifidobacterium longum JCM1217. Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 63, 751–753 (2007). doi: 10.1107/S1744309107036263 CrossRefPubMedGoogle Scholar
  23. 23.
    Suzuki, R., Wada, J., Katayama, T., Fushinobu, S., Wakagi, T., Shoun, H., et al.: Structural and thermodynamic analyses of solute-binding protein from Bifidobacterium longum specific for core 1 disaccharide and lacto-N-biose I. J. Biol. Chem. 283, 13165–13173 (2008). doi: 10.1074/jbc.M709777200 CrossRefPubMedGoogle Scholar
  24. 24.
    Kitaoka, M., Tian, J., Nishimoto, M.: Novel putative galactose operon involving lacto-N-biose phosphorylase in Bifidobacterium longum. Appl. Environ. Microbiol. 71, 3158–3162 (2005). doi: 10.1128/AEM.71.6.3158-3162.2005 CrossRefPubMedGoogle Scholar
  25. 25.
    Nishimoto, M., Kitaoka, M.: Identification of the putative proton donor residue of lacto-N-biose phosphorylase (EC 2.4.1.211). Biosci. Biotechnol. Biochem 71, 1587–1591 (2007). doi: 10.1271/bbb.70064 CrossRefPubMedGoogle Scholar
  26. 26.
    Hidaka, M., Nishimoto, M., Kitaoka, M., Wakagi, T., Shoun, H., Fushinobu, S.: The crystal structure of galacto-N-biose/lacto-N-biose I phosphorylase: a large deformation of a TIM barrel scaffold. J. Biol. Chem. 284, 7273–7283 (2009). doi: 10.1074/jbc.M808525200 CrossRefPubMedGoogle Scholar
  27. 27.
    Suzuki, R., Katayama, T., Ashida, H., Yamamoto, K., Kitaoka, M., Kumagai, H., et al.: Crystallographic and mutational analyses on the substrate specificity of endo-α-N-acetylgalactosaminidase from Bifidobacterium longum. J. Biochem. (Tokyo). in press. doi: 10.1093/jb/mvp086

Copyright information

© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Hisashi Ashida
    • 1
  • Hayato Ozawa
    • 1
  • Kiyotaka Fujita
    • 2
  • Shun’ichi Suzuki
    • 3
  • Kenji Yamamoto
    • 1
  1. 1.Graduate School of BiostudiesKyoto UniversityKyotoJapan
  2. 2.Faculty of AgricultureKagoshima UniversityKagoshimaJapan
  3. 3.Ajinomoto Co., Inc.KawasakiJapan

Personalised recommendations