Biochemical characterisation of extracellular phytase (myo-inositol hexakisphosphate phosphohydrolase) from a hyper-producing strain of Aspergillus niger van Teighem |
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Authors: | Purva Vats U. C. Banerjee |
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Affiliation: | (1) Institute of Microbial Technology, Sector 39 A, Chandigarh, 160 036, India;(2) Department of Pharmaceutical Technology, National Institute of Pharmaceutical Education and Research, Sector 67, SAS Nagar, 160 062, Punjab, India |
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Abstract: | Aspergillus niger van Teighem, isolated in our laboratory from samples of rotten wood logs, produced extracellular phytase having a high specific activity of 22,592 units (mg protein)–1 . The enzyme was purified to near homogeneity using ion-exchange and gel-filtration chromatography. The molecular properties of the purified enzyme suggested the native phytase to be oligomeric, with a molecular weight of 353 kDa, the monomer being 66 kDa. The purified enzyme exhibited maximum activity at pH 2.5 and 52–55°C. The enzyme retained 97% activity after a 24-h incubation at 55°C in the presence of 10 mM glycine, while 87% activity was retained when no thermoprotectant was added. Phytase activity was not affected by most metal ions, inhibitors and organic solvents. Non-ionic and cationic detergents (0.1–5%) stabilise the enzyme, while the anionic detergent (SDS), even at a 0.1% level, severely inhibited enzyme activity. The chaotropic agents guanidinium hydrochloride, urea, and potassium iodide (0.5–8 M), significantly affected phytase activity. The maximum hydrolysis rate (Vmax) and apparent Michaelis-Menten constant (Km) were 1,074 IU/mL and 606 M, respectively, with a catalytic turnover number of 3×105 s–1 and catalytic efficiency of 3.69×108 M–1 s–1. |
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Keywords: | Aspergillus niger Phytases Acid-phosphatases |
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