Graphical AbstractAs a natural sweetening and solubilizing agent, rubusoside has great potential in the application of healthy beverages and pharmaceuticals. A novel β-glucosidase CsBGL obtained here from Chryseobacterium scophthalmum 1433 through screening of the environmental microorganisms could specifically hydrolyze the C-13- or/and C-19-linked sophorese of steviol glycosides, but not the C-13- or/and C-19-linked Glcβ1-2[Glcβ1-3]Glc trisaccharide and Glcβ1-monosaccharide of steviol glycosides, and thus could be a promising biocatalyst for efficient rubusoside production from stevioside.

Sialidases are increasingly used in the production of sialyloligosaccharides, a significant component of human milk oligosaccharides. Elucidating the catalytic mechanism of sialidases is critical for the rational design of better biocatalysts, thereby facilitating the industrial production of sialyloligosaccharides. Through comparative all-atom molecular dynamics simulations, we investigated the structural dynamics of sialidases in Glycoside Hydrolase family 33 (GH33). Interestingly, several sialidases displayed significant conformational transition and formed a new cleft in the simulations. The new cleft was adjacent to the innate active site of the enzyme, which serves to accommodate the glycosyl acceptor. Furthermore, the residues involved in the specific interactions with the substrate were evolutionarily conserved in the whole GH33 family, highlighting their key roles in the catalysis of GH33 sialidases. Our results enriched the catalytic mechanism of GH33 sialidases, with potential implications in the rational design of sialidases.