Biocatalytic production of UDP-glucose from corn stover via DES pre-treatment

Current UDP-glucose production mainly depends on chemical synthesis, sucrose-based enzymatic routes, or intracellular microbial metabolism, which are limited by high cost, reliance on food-derived substrates, or complex pathway regulation. Direct production of UDP-glucose from non-food lignocellulosic biomass remains largely unexplored. In this study, a weak alkaline-assisted deep eutectic solvent pre-treatment was integrated with enzymatic hydrolysis and whole-cell biocatalysis to convert corn stover into UDP-glucose. ChCl/EG supplemented with 2% NaHCO3 achieved 81.08% lignin removal while retaining 73.99% cellulose under optimized conditions of 140 °C for 3 h at a solid–liquid ratio of 1:20. Structural characterization showed substantial surface disruption and cellulose enrichment, with the crystallinity index increasing from 39.24% to 64.99%. As a result, enzymatic hydrolysis was enhanced, giving 24.53 g/L glucose and 10.47 g/L xylose, approximately 6-fold and 3-fold higher than those obtained from untreated corn stover, respectively. A multi-enzyme system combining low-β-glucosidase cellulase with Escherichia coli surface-displayed cellobiose phosphorylase, UDP-glucose pyrophosphorylase, and polyphosphate kinase enabled UDP-glucose synthesis, reaching a maximum concentration of 7.13 mM. These results demonstrate an integrated non-food biomass-to-nucleotide-sugar route and provide a sustainable strategy for converting lignocellulose into high-value biochemical intermediates.