Mechanistic partitioning and functional synergy of bacterial cellulosome and fungal cellulases in cellulose saccharification.

Synergistic saccharification involving bacterial cellulosome and fungal cellulases holds significant promise for industrial applications; however, this integrated approach remains largely under-investigated. This study dissects the synergy from structural characterization, adsorption analysis, crystallinity evolution, and hydrolysis performance evaluation. The cellulosome exhibited superior enhancement of substrate accessibility after enzymatic hydrolysis of sugarcane bagasse compared with fungal cellulases. The distinct changes in crystallinity observed during the hydrolysis of microcrystalline cellulose by these two enzymes suggest differences in their adsorption affinities toward the crystalline and amorphous regions of cellulose. Further adsorption analysis demonstrated that fungal cellulases exhibited a higher adsorption capacity for amorphous cellulose (0.70 μmol/g). Cellulosome exhibited stronger apparent affinity toward crystalline cellulose (0.34 μmol/g vs. 0.24 μmol/g). Faster adsorption rates were observed on crystalline substrates, whereas higher equilibrium capacities occurred on amorphous regions. Apparent thermodynamic analysis confirmed spontaneous and exothermic adsorption. Apparent adsorption affinity followed: amorphous cellulose-fungal cellulase > amorphous cellulose-cellulosome > crystalline cellulose-cellulosome > crystalline cellulose-fungal cellulase. Optimal enzyme ratios should be tailored to substrate crystallinity to maximize synergistic saccharification efficiency. These results indicate that the observed synergy is governed by complementary substrate targeting and differential hydrolytic behavior, providing a rational basis for optimizing enzymatic systems.