Purification and biochemical characterization of an extracellular fructosyltransferase enzyme from Aspergillus niger sp. XOBP48: implication in fructooligosaccharide production.

An extracellular fructosyltransferase (Ftase) enzyme with a molar mass of ≈70 kDa from a newly isolated indigenous coprophilous fungus Aspergillus niger sp. XOBP48 is purified to homogeneity and characterized in this study. The enzyme was purified to 4.66-fold with a total yield of 15.53% and specific activity of 1219.17 U mg-1 of protein after a three-step procedure involving (NH4 )2 SO4 fractionation, dialysis and anion exchange chromatography. Ftase showed optimum activity at pH 6.0 and temperature 50 °C. Ftase exhibited over 80% residual activity at pH range of 4.0-10.0 and ≈90% residual activity at temperature range of 40-60 °C for 6 h. Metal ion inhibitors Hg2+ and Ag+ significantly inhibited Ftase activity at 1 mmol concentration. Ftase showed K m , v max and k cat values of 79.51 mmol, 45.04 µmol min-1 and 31.5 min-1 , respectively, with a catalytic efficiency ( k cat / K m ) of 396 µmol-1  min-1 for the substrate sucrose. HPLC-RI experiments identified the end products of fructosyltransferase activity as monomeric glucose, 1-kestose (GF2 ), and 1,1-kestotetraose (GF3 ). This study evaluates the feasibility of using this purified extracellular Ftase for the enzymatic synthesis of biofunctional fructooligosaccharides.