Characterization and kinetic parameters of endo-1,4-β-d-glucanase of ultraviolet mutated Aspergillus niger and Penicillium citrinum

Abstract

Mutagenic agents had been used to enhance production of enzymes in microorganisms, but the effects of the agents on the kinetic parameters and catalytic efficiency of the produced enzymes had not been critically evaluated. A study was conducted to assess the effects of ultraviolet (UV) mutagenesis on the kinetic parameters and catalytic efficiency of endo-1,4-β-D-glucanase enzyme obtained from wild and mutated strains of Aspergillus niger and Penicillium citrinum obtained from decaying wood wastes modified by ultraviolet radiation. The endo-1, 4-β-Dglucanase from both wild and mutant strains of the fungi were purified by centrifugation, ammonium sulfa-te precipitation and anion exchange chromatography using Sephadex A25-120 and Whatman DE-52 resins. The kinetic parameters determined from Lineweaver-Burk plot, using the velocity or reaction rates at varying concentrations of carboxymethyl cellulose substrate ranging from 0 - 20 mg/mL were validated by Hanes-Woolf plots for convergence, as the Michaelis-Menten kinetic equation is non-linear. The Michaelis-Menten constant (K_m) and maximum velocity (V_max) for the wild A. niger were 15.31 g/L and 4.67 g/L respectively, while values for corresponding mutated strain were 7.9 g/L and 3.39 g/L respectively. The K_m and V_max for wild P. citrinum were 7.94g/L and 6.81g/L respectively, while the values for the mutated strain were 6.60 g/L and 6.84 g/L respectively. Catalytic efficiency of endoglucanase of wild and mutated strains of A. niger were 0.305 and 0.42 M^-1s^-1 respectively, while the values for endoglucanase of wild and mutated strains of P. citrinum were 0.858 and 1.036 M^-1s^-1 respectively. Mutagenesis using ultraviolet radiation therefore enhanced the catalytic efficiency of endoglucanase enzyme.

Keywords: Endo-1,4-β-D-glucanase, Aspergillus niger, Penicillium citrinum, UV Mutagenesis, Michaelis-Menten Kinetic Equation, Lineweaver-Burk Plot, Hanes Woolf Plot, Catalytic Efficiency