An acidophilic GH12 xyloglucanase from Trichoderma asperellum produces prebiotic oligosaccharides that promote probiotic growth.

Enzymatic transformation of lignocellulosic biomass offers an eco-friendly pathway to produce valuable β-oligoglucosides. In this study, an acidophilic GH12 xyloglucanase (TaXEG12) from Trichoderma asperellum ND-1 was overexpressed in Komagataella phaffii. TaXEG12 displayed maximum activity at 40 °C and a pH of 4.0, while retaining >80% activity in the presence of 1.71 M NaCl and 10% ethanol. Glu140 and Glu227 are important catalytic residues, while Asp126 plays an auxiliary role in xyloglucan degradation. The enzyme showed strict substrate specificity toward tamarind xyloglucan. Through fed-batch fermentation, the enzyme production of TaXEG12 improved by 28.5-fold (2914.5 ± 65.7 U/mL). Interestingly, TLC and ESI-MS analyses revealed that TaXEG12 could hydrolyze xyloglucan into xyloglucan oligosaccharides (XXXG, XXLG/XLXG, and XLLG). These xyloglucan oligosaccharides can promote the growth of Lactobacillus bulgaricus. Additionally, the synergy degrees of corn bran and apple pulp in the degradation by TaXEG12 and TrCel were 1.34 and 1.32, respectively. TaXEG12 displayed unique properties and could be a desirable catalyst for the more sustainable production of high-value oligosaccharides, which are significantly beneficial for human intestinal health.