Enhancing the acid stability of the recombinant GH11 xylanase xynA through N-terminal substitution to facilitate its application in apple juice clarification.

The utilization of xylanase in juice clarification is contingent upon its stability within acidic environments. We generated a mutant xynA-1 by substituting the N-terminal segment of the recombinant xylanase xynA to investigate the correlation between the N-terminal region of xylanase and its acid stability. The enzymatic activity of xynA-1 was found to be superior under acidic conditions (pH 5.0). It exhibited enhanced acid stability, surpassing the residual enzyme activity values of xynA at pH 4.0 (53.07 %), pH 4.5 (69.8 %), and pH 5.0 (82.4 %), with values of 60.16 %, 77.74 %, and 87.3 %, respectively. Additionally, the catalytic efficiency of xynA was concurrently improved. Through molecular dynamics simulation, we observed that N-terminal shortening induced a reduction in motility across most regions of the protein structure while enhancing its stability, particularly Lys131-Phe146 and Leu176-Gly206. Furthermore, the application of treated xynA-1 in the process of apple juice clarification led to a significant increase in clarity within a short duration of 20 min at 35 °C while ensuring the quality of the apple juice. This study not only enhances the understanding of the N-terminal region of xylanase but also establishes a theoretical basis for augmenting xylanase resources employed in fruit juice clarification.