A thermophilic, pH-tolerant, and highly active GH10 xylanase from Aspergillus fumigatus boosted pre-treated sugarcane bagasse saccharification by cellulases

We have heterogously expressed a recombinant GH10 xylanase from Aspergillus fumigatus Af293 (Af-XYLA) in Pichia pastoris X-33 and investigated its biochemical and kinetic properties and its synergy with cellulases during lignocellulose sacchararification. Af-XYLA showed optimal activity at 80 degrees C and pH 5-7, and it withstood a temperature of 50 degrees C and pH 4-8 for 72 h with about 50 % and 80 % of its maximal activity, respectively, in the absence of substrate. Kinetic assays revealed that it presented the highest V-max (35,756 U mg(-1) at 50 degrees C and 88,082 U mg(-1 )at 80 degrees C) and k(cat)/K-M (15,456 mL mg(-1) s(-1) at 50 degrees C and 15,081 mL mg(-1) s(-1) at 80 degrees C) constants among fungal GH10 xylanases. Af-XYLA was xylose- and glucose-tolerant and displayed >70 % activity at 10 % (v/v) ethanol. Few additives affected Af-XYLA negatively, and the surfactants Tween 20, Triton X-100, and SLS and the reducing agents DTT and beta-mercaptoethanol improved enzyme performance by 28 %, 11 %, 8%, 44 %, and 32 %, respectively. We applied a 3-factor Central Composite Rotational Design (CCRD) to optimize important parameters for biomass saccharification and thus maximize the reducing sugars yields obtained from sugarcane exploded bagasse (SEB) and sugarcane delignified bagasse (SDB) hydrolyses. We carried out synergism assays under the previously determined optimal concentrations of Celluclast 1.5 (R) L (1.8 FPU g(-1)), Af-XYLA (3.75 mu g g(-1)), and Tween 20 (0.75 % v/v). Af-XYLA boosted SEB and SDB hydrolyses by cellulases by 155 % and 277 %, respectively, after 48 h. On the basis of the unique properties of Af-XYLA, it is a robust enzyme for application in biorefineries.

Automated summary

The recombinant GH10 xylanase Af-XYLA from Aspergillus fumigatus Af293 displayed optimal activity at 80 degrees C and pH 5-7, showing high tolerance to xylose, glucose, and ethanol. It exhibited excellent kinetic properties and was synergistic with cellulases, significantly boosting reducing sugars yields from biomass saccharification.