Statistical Optimization of Expression Cassettes Encoding 1, 4-?-D-Glucan Glucohydrolase from A Novel Strain Aspergillus Oryzae IIB-6

Background: Exo-1, 4-?-D-glucan glucohydrolase (GGH) is a biological catalyst that catalysis the starch saccharification to produce glucose syrup, which is used as feedstock for fuel ethanol and high fructose corn syrup production. Objective: The study aimed to optimize cost-effective GGH production from a novel strain IIB-6 by a statistical method. Methodology: The 18S rRNA sequence was used for the identification of soil isolate IIB-6. Different agricultural waste residues and moistening agents were evaluated for the production of GGH. The cumulative effects of wheat bran, moisture content, trace elements, and incubation time were optimized using a central composite rotatable design (CCRD) and response surface methodology (RSM). Result: The strain IIB-6 was identified as Aspergillus oryzae by phylogenetic analysis. Wheat bran 7.4 g, moisture contents 67 %, trace elements (mg.gds-1) KH2PO4 0.402; K2HPO4 0.938; MgSO4.7H2O 0.67; (NH4)2SO4 2.68; FeSO4.7H2O 0.0134, and time of incubation 86 h were the best culture conditions for the GGH production. Expression cassettes GGH1, GGH2, and GGH3 were isolated and purified. Conclusion: It is concluded that the statistical optimization method is highly effective in accommodating even minor variations in culture conditions to yield reliable results. The novel strain Aspergillus oryzae IIB-6 is a promising source of GGH due to the high enzyme titer released, which encouraged the technologists to isolate expression cassettes encoding for GGH, for further implementation in the industrial strain development processes.