Abstract
Xyloglucan, a key component of plant cell wall polysaccharides, plays a crucial role in cell wall structural remodeling and biomass recalcitrance. This study reports the discovery and biochemical characterization of a novel glycoside hydrolase family 12 (GH12) xyloglucanase, TpXEG12a, from the biomass-degrading fungus Talaromyces pinophilus. Recombinant TpXEG12a exhibited exceptional catalytic efficiency toward xyloglucan, with a specific activity of 2375 U/mg, significantly higher than the typical range reported for GH12 xyloglucanases. The enzyme displayed optimal activity at pH 4.0 and 57 °C, with high stability in acidic conditions (pH 4-8) and moderate thermal stability. TpXEG12a demonstrated strict substrate specificity for xyloglucan, with no detectable activity against cellulose-related substrates, and primarily generated characteristic xyloglucan oligosaccharides (XXXG, XLXG/XXLG, XLLG) upon hydrolysis. Structural analysis revealed that TpXEG12a exists as a stable homodimer in solution, which likely contributes to its catalytic efficiency. Notably, TpXEG12a synergistically enhanced glucose release when combined with cellulase in lignocellulosic biomass degradation. These findings establish TpXEG12a as a promising candidate for industrial applications in biomass conversion, textile processing, and functional oligosaccharide production.