The Importance of Oligosaccharide Synthesis

Oligosaccharide synthesis is becoming increasingly important to the pharmaceutical industry because of the great potential of these complex molecules as therapeutic agents. Furthermore, some oligosaccharides are used as additives in food ingredients because they exhibit beneficial effects by stimulating the growth and/or activity of gut bacteria.

Due to the excellent regioselectivity of certain catalysts, no orthogonal protection/deprotection of different moieties is required and the control over the stereochemistry of the new bonds formed is complete. There are two main methods for enzymatic oligosaccharide synthesis: glycosyltransferases and glycosidases. Glycosyltransferases are enzymes responsible for the biosynthesis of oligosaccharides in vivo. In this case, the sugar must be activated at the anomeric center by nucleoside phosphate or monophosphate. Glycosidases, on the other hand, belong to the family of hydrolases whose natural function is to cleave glycosidic bonds. Glycosidases catalyze reverse hydrolysis or transglycosylation with retention mechanisms.

Fig.1 Fructooligosaccharide production from sucrose by transfructosylation. (Mano, et al., 2018)

Enzymatic Transglycosylation for Oligosaccharide Manufacture at CD BioGlyco

CD BioGlyco has well-trained technicians and provides clients with comprehensive and reliable One-Stop Solutions for Carbohydrate Manufacture. We aspire to become the trusted solution provider of your first choice. We offer enzymatic transglycosylation for oligosaccharide manufacture using glycosidase. Glycosidases catalyze reverse hydrolysis or transglycosylation with retention mechanisms. When disaccharides are used as substrates, transglycosylated products are formed by self-condensation. Using cellobiose and/or maltose as substrates, we provide the following scheme for the preparation of glucosyl oligosaccharides for reference.

• Enzyme purification and activity assay

β-Glucosidase is obtained from strain culture or commercial sources. If the enzyme is from a cultured strain, a two-step protocol for purification is used. Extracts are first heat-treated, followed by affinity chromatography on immobilized metal ions using the supernatant from the heat-treatment step. Purified enzymes are stored at lower temperatures until use.

Enzyme activity assays are then performed by standard methods.

• Transglycosylation studies

Select the conditions under which the glycosidase exhibits higher activity including pH value, substrate concentration, and temperature conditions to experiment. The substrate sugar is dissolved and incubated for a while. Then, add glucosidase and shake the reaction at the optimum temperature. Finally, the enzymatic reaction is quenched by adding NaOH.

• Measurement of transglycosylation rate

The concentrations of different substances in the mixture are determined by high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD). The obtained products are qualitatively and quantitatively prepared by standard music. Initial rates are calculated by monitoring product concentrations over time.

Fig.2 The scheme for the preparation of glucosyl oligosaccharides. (CD BioGlyco)

Applications

• Oligosaccharides produced by enzymatic transglycosylation are important to the pharmaceutical industry.
• Oligosaccharides produced by enzymatic transglycosylation are used as food additives because they stimulate the growth of intestinal bacteria.
• The stereoselective and regioselective properties of enzyme catalysis are important advantages for Oligosaccharide Production and are complementary tools to chemical methods.

Advantages

• The catalysts are outstandingly regioselective so that orthogonal protection or deprotection of different groups is not required, and the stereochemical control over the new bonds formed is complete.
• Glycosidases catalyze dehydrolysis (thermodynamic control) or transglycosylation (kinetic control) under a retention mechanism.
• The Enzymatic Process for Oligosaccharide Manufacture is environmentally friendly.
• The rapid development of enzyme biology contributes to the progress of enzymatic transglycosylation to produce oligosaccharides.

At CD BioGlyco, we pay attention to every detail in enzymatic transglycosylation for oligosaccharide manufacture and strive for perfection in every process. Should you have any questions, please don't hesitate to reach out and contact us.

• Enzymatic Degradation/Hydrolysis for Oligosaccharide Manufacture
• Enzymatic Synthesis for Oligosaccharide Manufacture
• Enzymatic Isomerization for Oligosaccharide Manufacture
• Enzymatic Epimerization for Oligosaccharide Manufacture