title: Purification and characterization of an extracellular keratinolytic protease from a new isolate of Aspergillus parasiticus.
abstract: Keratinolytic proteases find extensive applications both in environmental biotechnology and pharmaceutical industries. An extracellular keratinolytic protease was purified and characterized from the fungus, Aspergillus parasiticus, isolated from poultry soil. The enzyme was purified to homogeneity by acetone and ammonium sulfate precipitations followed by CM-Sepharose column chromatography. The molecular mass of the enzyme was 36kDa as judged by SDS-PAGE. The purified keratinase had a pH optimum of 7.0 and temperature optimum of 50(o)C. The enzyme hydrolyzed the substrate azocasein and the Km and Vmax of the purified keratinase were found to be 1.04mg/ml and 3463.34Units/min/mg protein, respectively. The enzyme showed increased activity in the presence of reducing agents. The enzyme was found to be glycosylated. According to the inhibition profiles obtained with the various protease inhibitors, it was confirmed that the purified keratinase belongs to the serine protease type. The purified enzyme activity was enhanced by calcium, magnesium and manganese ions and partially inhibited by cadmium, copper and zinc ions. The purified enzyme showed increased activity with nonionic detergents and urea.
authors: Department of Molecular Microbiology, School of Biotechnology, Madurai Kamaraj University, Madurai - 625 021, Tamil Nadu, India.
authors: T S
authors: TS
authors: Anitha
authors: P
authors: P
authors: Palanivelu
mesh_terms: D000595:Amino Acid Sequence; D000645:Ammonium Sulfate / Q000737:chemistry; D001230:Aspergillus / Q000737:chemistry / Q000201:enzymology* / Q000302:isolation & purification; D011232:Chemical Precipitation; D003902:Detergents / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D019163:Reducing Agents / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013696:Temperature
chemical_list: D003902:Detergents; D019163:Reducing Agents; D010447:Peptide Hydrolases; C035855:keratinase; D000645:Ammonium Sulfate