Efficient transfer of sialo-oligosaccharide onto proteins by combined use of a glycosynthase-like mutant of Mucor hiemalis endoglycosidase and synthetic sialo-complex-type sugar oxazoline

Midori Umekawa, Takayuki Higashiyama, Yurie Koga, Tomonari Tanaka, Masato Noguchi, Atsushi Kobayashi, Shin ichiro Shoda, Wei Huang, Lai Xi Wang, Hisashi Ashida, Kenji Yamamoto

Research output: Contribution to journalArticlepeer-review

71 Citations (Scopus)

Abstract

Background: An efficient method for synthesizing homogenous glycoproteins is essential for elucidating the structural and functional roles of glycans of glycoproteins. We have focused on the transglycosylation activity of endo-β- N-acetylglucosaminidase from Mucor hiemalis (Endo-M) as a tool for glycoconjugate syntheses, since it can transfer en bloc the oligosaccharide of not only high-mannose type but also complex-type N-glycan onto various acceptors having an N-acetylglucosamine residue. However, there are two major bottlenecks for its practical application: the low yield of the transglycosylation product and the difficulty to obtain the activated sugar oxazoline substrate, especially the sialo-complex type one. Methods: We carried out the transglycosylation using a glycosynthase-like N175Q mutant of Endo-M, which was found to possess enhanced transglycosylation activity with sugar oxazoline as a donor substrate, in combination with an easy preparation of the sialo-complex-type sugar oxazoline from natural sialoglycopeptide in egg yolk. Results: Endo-M-N175Q showed efficient transglycosylation toward sialo-complex-type sugar oxazoline onto bioactive peptides and bovine ribonuclease B, and each sialylated compound was obtained in significantly high yield. Conclusions: Highly efficient and simple chemo-enzymatic syntheses of various sialylated compounds were enabled, by a combination of a simple synthesis of sialo-complex-type sugar oxazoline and the Endo-M-N175Q catalyzed transglycosylation. General significance: Our method would be very useful for a practical synthesis of biologically important glycopeptides and glycoproteins.

Original languageEnglish
Pages (from-to)1203-1209
Number of pages7
JournalBiochimica et Biophysica Acta - General Subjects
Volume1800
Issue number11
DOIs
Publication statusPublished - 2010 Nov

Keywords

  • Chemo-enzymatic synthesis of glycoprotein
  • Glycosynthase-like mutant
  • Sialo-complex-type glycan
  • Sugar oxazoline
  • Transglycosylation

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

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