title: Purification, immobilization and characterization of tannase from Penicillium variable.
abstract: Tannase from Penicillium variable IARI 2031 was purified by a two-step purification strategy comprising of ultra-filtration using 100 kDa molecular weight cutoff and gel-filtration using Sephadex G-200. A purification fold of 135 with 91% yield of tannase was obtained. The enzyme has temperature and pH optima of 50 degrees C and 5 degrees C, respectively. However, the functional temperature range is from 25 to 80 degrees C and functional pH range is from 3.0 to 8.0. This tannase could successfully be immobilized on Amberlite IR where it retains about 85% of the initial catalytic activity even after ninth cycle of its use. Based on the Michaelis-Menten constant (Km) of tannase, tannic acid is the best substrate with Km of 32 mM and Vmax of 1.11 micromol ml(-1)min(-1). Tannase is inhibited by phenyl methyl sulphonyl fluoride (PMSF) and N-ethylmaleimide retaining only 28.1% and 19% residual activity indicating that this enzyme belongs to the class of serine hydrolases. Tannase in both crude and crude lyophilized forms is stable for one year retaining more than 60% residual activity.
authors.affiliation: Department of Microbiology, University of Delhi South Campus, Benito Juarez Road, New Delhi 110 021, India.
authors.forename: Shashi
authors.initials: S
authors.lastname: Sharma
authors.forename: Lata
authors.initials: L
authors.lastname: Agarwal
authors.forename: Rajendra Kumar
authors.initials: RK
authors.lastname: Saxena
mesh_terms: D002265:Carboxylic Ester Hydrolases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D010407:Penicillium / Q000201:enzymology*; D013379:Substrate Specificity
chemical_list: D004791:Enzyme Inhibitors; D004800:Enzymes, Immobilized; D002265:Carboxylic Ester Hydrolases; C022711:tannase