Gene cloning and characterization of an organic solvent-stimulated β-glucosidase and its application for the co-production of ethanol and succinic acid

Enzymatic activity is a limiting factor for conducting the hydrolysis of lignocellulosic waste materials to generate renewable biofuels and biochemicals. A novel beta-glucosidase (BGL) gene from Hypocrea sp. W-63 was cloned and expressed in Pichia pastoris. The specific activity of the purified recombinant epB-BGL was 194.25 IU/mg using p-nitrophenyl-beta-D-glucoside (pNPG) as a substrate. The optimum pH and temperature for epB-BGL were 5.0 and 70 degrees C, respectively. The activity of recombinant epB-BGL could be stimulated by 210% with the addition of 10% (v/v) ethanol. Furthermore, other organic solvents were also able to stimulate the activity of the enzyme at concentrations of 1% or 10% (v/v). Due to this distinctive performance, epB-BGL was used to release fermentable sugars from a sugarcane bagasse hydrolysate for ethanol and succinic acid co-production. In the experimental range, succinic acid was reached to 13.3 g/L with a higher yield of 0.91 g/g glucose and the ethanol was at a concentration of approximately 1.41 g/L, respectively. In addition, the by-product acetate, which could potentially be converted to other chemicals, was also produced at 9.745 g/L. This work demonstrates that recombinant epB-BGL can take advantage of incomplete hydrolysis to convert fermentable sugars, suggesting its potential application in lignocellulose bioconversion.