Comparative kinetics evidence of a novel 1, 4-α-D-glucan glucohydrolase from Aspergillus oryzae IIB-6 as a raw starch-degrading enzyme

Raw starch-digesting enzymes (RSDEs) have industrial applications, where they eliminate the need for pre-gelatinization/-liquefaction in glucose syrup production. This syrup is then used as feedstock for cost-effective manufacturing of various value-added industrial products, including biofuels, biochemicals, biomaterials, and pharmaceutical products. The study aimed to produce a novel raw starch-digesting 1,4-α-D-glucan glucohydrolase (RSD-GGH) from a GRAS strain, Aspergillus oryzae IIB-6, via solid-state fermentation. Partially purified RSD-GGH at 50-70% (NH4)2SO4 saturation was used to saccharify raw starches extracted from different botanical sources, to evaluate their extent (Rs) and degree of hydrolysis (DE, %) compared to soluble starch. The amylose (AM) and amylopectin (AMP) contents of raw starches from rice, wheat, maize, sweet potato, and potato were 16.56, 20.53, 25.13, 30.56, and 51.65 % (LSD ~0.753), and 83.44, 79.47, 74.87, 69.44, and 48.37 % (LSD~1.202), respectively. The Rs and DE values of potato-based soluble, rice, wheat, corn, sweet potato, and potato raw starches were calculated as 3.75, 2.65, 1.6, 0.7, 0.3, 0.1 % and 0.0375, 0.0265, 0.016, 0.007, 0.003, and 0.001 %, respectively, while the Km and Vmax values at 60 °C, pH 5, were 14.851, 36.875, 40.671, 57.192, 113.23, and 453.01 mg.ml-1, and 34.488, 1250.2, 833.34, 476.19, 370.37, and 454.02 mg.ml-1.min-1, respectively. The extent of saccharification (Rs) and dextrose equivalent (DE) values were also calculated. Significant variations (p ≤ 0.05) in AM and AMP contents, as well as the botanical origin of the raw starches, influenced the enzyme’s kinetics accordingly. The enzyme has great potential for the industrial production of fuel ethanol and other biotechnological products. Keywords: Raw starch digesting enzyme, 1, 4-α-D-glucan glucohydrolase, raw starch extraction, dextrose equivalent, extent of saccharification, amylose, amylopectin, enzyme kinetics.