Bioprocessing and characterization of thermostable phytase from Aspergillus terreus, an endophyte of Catharanthus roseus, with a potential activity to hydrolyze phytic acid in wheat bran.

Phytic acid is one of the common anti-nutritional factors in animal feeds, due to its chelating activity of metal ions and amino acids, so, phytase has been used for increasing the nutritional value of the animal feeds by releasing phosphorous. The stability and catalytic efficiency of this enzyme are the major challenges, so, the objective of this study was to purify and characterize phytase with relatively unique biochemical properties. Among the recovered fungal endophytes of Catharanthus roseus, Aspergillus terreus EFBL-AS PV412881.1 was recognized as the most potent phytase producing isolate. Upon nutritional optimization with the face-centered central composite design (FCCD), the productivity of phytase by A. terreus grown on wheat bran amended with 0.2% NaNO₃ and 0.4 % yeast extract, under solid state fermentation, was increased into 36.3 μmol/mg/min. Phytase of A. terreus was purified to its molecular homogeneity by gel-filtration and ion-exchange chromatography, with 3.48 purification folds (125 μmol/mg/min). The purified enzyme had a molecular subunit 85 kDa by denaturing-PAGE, with highest activity at reaction temperature 37-40 °C, and reaction pH 7.0. The T_1/2 of A. terreus phytase was 124.5, 5.2 and 3.8 h, at 4, 40, and 50 °C, respectively. The thermal denaturation rate (Kr) was 0.095 ×10^-3, 0.27 × 10^-3 and 0.292 ×10^-3/min at 20, 40, and 50 °C, respectively. The enzyme was slightly inhibited by Ca²⁺ ions, unlike the resistance to various cations. The concentration of phytic acid of wheat bran was reduced by about 6.5 folds upon phytase treatment, ensuring the feasibility of this enzyme in the animal feed application. From the molecular docking analysis, phytase from A. terreus had a higher affinity to hydrolyze phytic acid, with binding energy - 7.1 kcal/mol, compared to that of A. niger and P. pinophilum (-6.7 kcal/mol), ensuring the stability of the interaction.