REGULATION OF EXPRESSION OF MULTIPLE BETA- GLUCOSIDASES OF ASPERGILLUS TERREUS AND THEIR PURIFICATION AND CHARACTERIZATION

Abstract

This study reports the regulation and purification of -glucosidases from a thermotolerant Aspergillus terreus AN1 strain, previously reported for efficient deinking of composite paper waste. The differential expression of four -glucosidase isoforms, in response to carbon sources in production medium, was studied by electrophoretically resolving proteins by polyacrylamide gel electro-phoresis analysis (PAGE) and developing zymograms using methylum-belliferyl -D glucoside as substrate. Three -glucosidases (GI, GII & GIII) were purified using chromatographic techniques. SDS-PAGE revealed the respective molecular masses of GI, GII, and GIII, as 29, 43, and 98 KDa, and isoelectric point (pI) to be 2.8, 3.7, and 3.0. The -glucosidases exhibited diverse pH and temperature optima as well as stability. -Glucosidase I (GI) specifically recog-nized pNP--glucopyranoside (pNPG) as a substrate, whereas, -glucosidase II (GII) and III (GIII) also showed activities against cellobiose and salicin. In contrast to GII and GIII, the activity of GI was positively influenced in the presence of hexoses/pentoses and alcohols. Km and Vmax for hydrolysis of pNPG by GI, GII, andGIII were found to be 14.2 mM and 166.9 µmol -1mg protein -1, 4.37 mM, and 34.7 µmol -1mg proteins -1, and 11.1 mM and 378.7µ mol -1 mg protein -1, respectively.

Keywords

• Aspergillus terreus
• beta-glucosidases
• Multiplicity
• Differential expression
• Regulation
• Purification and characterization