Cloning, Overexpression, and Characterization of a Highly Active Endoinulinase Gene from Aspergillus fumigatus Cl1 for Production of Inulo-Oligosaccharides

In this study, an endoinulinase gene from Aspergillus fumigatus was cloned and overexpressed in Pichia pastoris. A maximum activity, 3860 U/ml, of the recombinant endoinulinase was obtained by using a high cell-density fermentation approach. The recombinant endoinulinase with a molecular weight of 58 kDa was purified for further enzymatic investigation. The pH and temperature optima were pH 6.0 and 55 A degrees C, and the K (m) and V (max) values toward inulin were 2.18 mM and 1590 mu mol min(-1) mg(-1), respectively. The degree of polymerization (DP) for inulo-oligosaccharides product 3, 4, 5, and > 5 was determined by thin-layer chromatography (TLC) and high-performance liquid chromatograph (HPLC). Immobilized endoinulinases were prepared and characterized, which showed higher stability than the free endoinulinase under various temperature levels. A residual activity of 81.2 % could be still obtained after ten reaction cycles. Thus, the present recombinant endoinulinase exhibited great potential for scale-up production of inulo-oligosaccharides.