A high-yield xylanase platform for efficient xylooligosaccharides production from agricultural residues

Agricultural residues are promising lignocellulosic feedstocks for producing xylooligosaccharides (XOS) through xylanase-mediated enzymatic hydrolysis. Achieving high XOS yield with cost-effective pretreatment and minimal enzyme loading is essential in biorefinery. This study achieved high XOS yield through enzymatic hydrolysis of agricultural residues. A robust platform for xylanase production was established using genetically engineered Pichia pastoris to secrete thermostable xylanase (tXyn2 from Trichoderma reesei). N-glycosylation of tXyn2 reduced its adsorption strength to lignin by 43 %. Through expression cassette optimization and chaperone co-expression, the expression of tXyn2 was enhanced to 10.2 g/L in a 5-L bioreactor, representing the highest reported secretory expression level for T. reesei xylanase. The saccharification of alkaline-pretreated corncob and cottonseed hull with optimized enzyme loading resulted in the XOS yields of 65.3 % and 77.5 %, respectively. The enzyme cost of XOS production from corncob is as low as 50 USD/ton XOS. Membrane-purified XOS exhibited significant growth-promoting effects on cucumber seedlings. Foliar application of XOS improved photosynthetic efficiency in cucumber seedlings by 51.4 % and elevated the indole-3-acetic acid content of roots by 45.6 %, which enhanced shoot height by 32.9 % and root biomass by 60.3 %. This study provides a cost-effective solution for XOS production and expands its application in agriculture.