Abstract:To further enhance the secretion and expression of xylanase (Xylanase, XynB), the locus of glucoamylase（Glucoamylase, GlaA）with endogenous high-expression of Aspergillus niger was utilized as the target site for gene integration. Based on the strong promoter of the six copies of the glaA gene, the expression and secretion of xynB were regulated by the signal peptide SglaA from glucoamylase and the signal peptide SpepB from acid protease B (Acid Protease B, PepB), respectively. Additionally, the impact of deleting the high-background protein acid-stable alpha-amylase （Acid-stable alpha-amylase, AsAA）on XynB secretion was also investigated. The results indicated that the recombinant strains TgX, TpX, ΔAgX, and ΔApX all displayed distinct XynB protein bands at approximately 22 kDa. Notably, the xylanase activity of strain ΔApX (21413.74 U/mL) was elevated by 70.67%, 14.92%, and 35.42% compared to strains TgX, TpX, and ΔAgX, respectively. The signal peptide exerts no significant effect on the transcription levels of the xynB gene and UPR marker genes. The absence of high-background protein AsAA can enhance the transcription level of the xynB gene and facilitate the folding and secretion of the recombinant protein, threrby alleviating endoplasmic reticulum stress. In summary, this study successfully optimized and enhanced the expression of XynB in Aspergillus niger through the utilization of efficient secretion signal peptides and the deletion of high-background proteins, thereby establishing a foundation for further continuous improvement of XynB.
