Stability and Thermodynamic Attributes of Starch Hydrolyzing α‐Amylase of Anoxybacillus rupiensis TS‐4

The physicochemical conditions influencing α‐amylase secretion by a thermophilic bacterium, Anoxybacillus rupiensis TS‐4 (Genbank number, KU360725) are optimized by the response surface methodology, using Plackett Burman design, followed by Box—Behnken design to enhance amylase production by threefold as compared to the one variable at a time approach. The amylase is purified by ion exchange chromatography, followed by size exclusion chromatography with fold purification and yield of 17.85% and 34.72%, respectively. The molecular weight, Km, and Vmax of the purified amylase are 48 kDa, 0.58 mg mL−1, and 3124 µmol mL−1 min−1, respectively. It catalyzes starch over a broader range of temperature and pH, having optima as 80 °C and 8, respectively. The enzyme is stable at a broad range of temperatures and pH, displaying higher half‐life and reduced deactivation rate constant. The feasibility of the starch catalysis reaction mediated by the studied amylase is substantiated by determining the thermodynamic parameters, such as alterations in the enthalpy, entropy, activation energy, and Gibb's free energy. The attributes of the amylase such as calcium independence, alkali‐tolerance, and stability in presence of various chelators and surfactants aid uniqueness, novelty, and commercial promise. The present study is focused on optimization of physiochemical parameters affecting production of α‐amylase by a thermophilic bacterium, Anoxybacillus rupiensis TS‐4 using response surface methodology, yielding almost fold‐fold enhanced production. Various thermodynamic parameters are determined to establish stability in the enzyme‐substrate reactions under range of extremities. Moreover, its possible commercial value is explored by calculating dextrose equivalent value (in production of high maltose containing syrups) and washing efficiencies (in the detergent industry).