Metabolic engineering and thermodynamic characterization of an extracellular β-glucosidase produced by Aspergillus niger

The production of an extracellular beta-glucosidase by Aspergillus niger NRRL 599 was optimized using submerged fermentation technique. Effect of different media, different carbon sources, initial pH of the fermentation medium, temperature, incubation period and inoculum size on the production of beta-glucosidase enzyme was investigated. A. niger NRRL 599 produced maximum extracellular beta-glucosidase (4.48 U/mg) in Eggins and Pugh medium with 1% wheat bran (w/v) at pH 5.5 inoculated with 4% conidial suspension after 96 h of incubation at 30 degrees C. Purified beta-glucosidase gave K-m and V-max values of 3.11 mM and 20.83 U/mg respectively for pNPG hydrolysis. The enzyme was optimally active at pH 4.8 and at temperature of 60 degrees C. Thermodynamic parameters, E-a, Delta H and Delta S were found to be 52.17 KJ/mol, 49.90 J/mol.K and -71.69 KJ/mol, respectively. The pKa(1) and pKa(2) of ionizable groups of active site residues involved in V-max were calculated to be 4.1 and 6.0 respectively.