Evolving trend of Boletus versicolor IBL-04 by chemical mutagenesis to overproduce laccase: Process optimization, 3-step purification, and characterization

Abstract

In this study, Boletus versicolor IBL-04 was exploited to produce laccase in solid-state fermentation of wheat straw. Wild-type strain of Boletus versicolor (Bv IBL-04) produced laccase with maximum activity of 118.89 ± 11.32 U/mL, which was further improved by manipulating the fungal genome through random chemical mutagenesis to gain a maximum enzyme productivity of 403.34 ± 13.79 U/mL. The maximum laccase-producing mutant strain (Bv EB-75ʺ) was selected by the double-screening process and proceeded to optimize the culture conditions for laccase production using response surface methodology (RSM). Through optimization, laccase activity was improved to 695.78 ± 6.7 U/mL, indicating a 3-fold relative increase in the production yield. The optimally produced enzyme fraction from mutant isolates was eventually purified using 3-step purification system of: (i) (NH4)2SO4 precipitation; (ii) ion-exchange chromatography and (iii) gel-filtration. By purification, a final purification fold of 44.07 was obtained with total enzyme and laccase specific activities of 9406 ± 15.094 IU/mL and 855.09 ± 22.065 U/mg, respectively. Characterization studies revealed the comparative improvement in pH and temperature profiles of laccases from mutant isolates than the wild-type strain. In conclusion, the mutagenesis improved the stability of laccase at higher pH and temperature values, indicating BvEB-75ʺ a potent fungal strain for laccases production for various biotechnological applications.