Three-Enzyme Phosphorylase Cascade for Integrated Production of Short-Chain Cellodextrins

Cellodextrins are linear β-1,4-gluco-oligosaccharides that are soluble in water up to a degree of polymerization (DP) of ≈6. Soluble cellodextrins have promising applications as nutritional ingredients. A DP-controlled, bottom-up synthesis from expedient substrates is desired for their bulk production. Here, a three-enzyme glycoside phosphorylase cascade is developed for the conversion of sucrose and glucose into short-chain (soluble) cellodextrins (DP range 3-6). The cascade reaction involves iterative β-1,4-glucosylation of glucose from α-glucose 1-phosphate (αGlc1-P) donor that is formed in situ from sucrose and phosphate. With final concentration and yield of the soluble cellodextrins set as targets for biocatalytic synthesis, three major factors of reaction efficiency are identified and partly optimized: the ratio of enzyme activity, the ratio of sucrose and glucose, and the phosphate concentration used. The efficient use of the phosphate/αGlc1-P shuttle for cellodextrin production is demonstrated and the soluble product at 40 g L^-1 is obtained under near-complete utilization of the donor substrate offered (88 mol% from 200 mm sucrose). The productivity is 16 g (L h)^-1 . Through a simple two-step route, the soluble cellodextrins are recovered from the reaction mixture in ≥95% purity and ≈92% yield. Overall, this study provides the basis for their integrated production.