Production, characterisation and applications of a thermo-acid-stable phytase from Enterobacter sp. ACSS
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A bacterial strain producing an extracellular phytase was identified as Enterobacter sp. ACSS. Optimization of process parameters using statistical methods such as Plackett-Burman design (PBD), the steepest ascent method, and response surface methodology (RSM) significantly improved phytase production by 4.6–fold in shake-flasks. In addition, an overall 1.9-fold increase in phytase production was attained in fed-batch fermentations in a 5 l laboratory fermenter, respectively. The purified 62 kDa phytase from Enterobacter sp. ACSS was active between 40 to 80°C and an acidic pH range of 2.0 to 6.0 with half-life of 693 and 577.5 min at 60°C and pH 2.0, respectively. Additionally, the enzyme is fairly stable with proteolytic enzymes under physiological conditions. It was activated by Ca+2, Mg+2 and Mn+2 while inhibition was caused by Zn+2, Cu+2, Fe+2, Pb+2, Co+2, Ba+2 and surfactants. The Km, Vmax and Kcat observed were 0.21 mM, 131.58 nmol mg-1s-1 and 1.64 × 103 s-1, respectively. The enzyme released inorganic phosphate from animal feed (4.0-6.62 mg/g of diet) and insoluble metal-phytates (45-219 µg/ml) and was effective in improving the characteristics of brown bread. Overall, this study shows that Enterobacter sp. ACSS has the potential to produce significant titres of a thermo- and acid-stable phytase and can be applied in dephytinizing animal feeds, and the baking industry.