Coimmobilization and colocalization of a glycosyltransferase and a sucrose synthase greatly improves the recycling of UDP-glucose: Glycosylation of resveratrol 3-O-ß-D-glucoside.

Glycosylation is one of the most efficient biocompatible methodologies to enhance the water solubility of natural products, and therefore their bioavailability. The excellent regio- and stereoselectivity of nucleotide sugar-dependent glycosyltransferases enables single-step glycosylations at specific positions of a broad variety of acceptor molecules without the requirement of protection/deprotection steps. However, the need for stoichiometric quantities of high-cost substrates, UDP-sugars, is a limiting factor for its use at an industrial scale. To overcome this challenge, here we report tailor-made coimmobilization and colocalization procedures to assemble a bi-enzymatic cascade composed of a glycosyltransferase and a sucrose synthase for the regioselective 5-O-ß-D-glycosylation of piceid with in situ cofactor regeneration. Coimmobilization and colocalization of enzymes was achieved by performing slow immobilization of both enzymes inside the porous support. The colocalization of both enzymes within the porous structure of a solid support promoted an increase in the overall stability of the bi-enzymatic system and improved 50-fold the efficiency of piceid glycosylation compared with the non-colocalized biocatalyst. Finally, piceid conversion to resveratrol 3,5-diglucoside was over 90% after 6 cycles using the optimal biocatalyst and was reused in up to 10 batch reaction cycles accumulating a TTN of 91.7 for the UDP recycling.

Designing continuous flow reaction of xylan hydrolysis for xylooligosaccharides production in packed-bed reactors using xylanase immobilized on methacrylic polymer-based supports.

The present study focuses on the development and optimization of a packed-bed reactor (PBR) for continuous production of xylooligosaccharides (XOS) from xylan. For this purpose, three different methacrylic polymer-based supports (Relizyme R403/S, Purolite P8204F and Purolite P8215F) activated with glyoxyl groups were morphologically characterized and screened for the multipoint covalent immobilization of a xylanase. Based on its physical and mechanical properties, maximum protein loading and thermal stability, Relizyme R403/S was selected to set up a PRB for continuous production of XOS from corncob xylan. The specific productivity for XOS at 10 mL/min flow rate was 3277 gXOS genzyme-1 h-1 with a PBR. This PBR conserved >90% of its initial activity after 120 h of continuous operation.