Artificial mixed-linked beta-glucans produced by glycosynthase-catalyzed polymerization: tuning morphology and degree of polymerization

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Abstract

The glycosynthase derived from Bacillus licheniformis 1,3-1,4-β- glucanase was able to polymerize glycosyl fluoride donors (G4) mG3GαF (m = 0-2, G = Glcβ) leading to artificial mixed-linked β-glucans with regular sequences and variable β1,3 to β1,4 linkage ratios. With the E134A glycosynthase mutant, polymers had average molecular masses (Mw) of 10-15 kDa. Whereas polymer 2 ([4G4G3G]n) was an amorphous precipitate, the water-insoluble polymers 1 ([4G3G]n) and 3 ([4G4G4G3G]n) formed spherulites of 10-20 μm diameter. With the more active E134S glycosynthase mutant, polymerization led to high molecular mass polysaccharides, where M w was linearly dependent on enzyme concentration. Remarkably, a homo-polysaccharide [4G4G4G3G]n with Mw as high as 30.5 kDa (n ≃ 47) was obtained, which contained a small fraction of products up to 70 kDa, a value that is in the range of the molecular masses of low viscosity cereal 1,3-1,4-β-glucans, and among the largest products produced by a glycosynthase. Access to a range of novel tailor-made β-glucans through the glycosynthase technology will allow to evaluate the implications of polysaccharide fine structures in their physicochemical properties and their applications as biomaterials, as well as to provide valuable tools for biochemical characterization of β-glucan degrading enzymes and binding modules.

Original languageEnglish
Pages (from-to)494-501
Number of pages8
JournalBiomacromolecules
Volume12
Issue number2
DOIs
Publication statusPublished - Feb 2011

Keywords

  • Xyloglucan endo-transglycosylase
  • Site-directed mutagenesis
  • In-vitro synthesis
  • Oligosaccharide synthesis
  • Bacillus-licheniformis
  • 1,3-1,4-beta-d-glucan 4-glucanohydrolases
  • Efficient synthesis
  • Mutant cellulase
  • Glycosidases
  • Glucosidase

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