Glucose production from cellulose through biological simultaneous enzyme production and saccharification using recombinant bacteria expressing the β-glucosidase gene

Abstract

Efficient cellulosic biomass saccharification technologies are required to meet biorefinery standards. Biological simultaneous enzyme production and saccharification (BSES), which is glucose production from cellulosic biomass by Clostridium thermocellum, can be a reliable cellulose saccharification technology for biorefineries. However, the current BSES processes require purified β-glucosidase supplementation. In this study, recombinant bacteria expressing the β-glucosidase gene were developed and directly applied to BSES. The engineered Escherichia coli expressing the thermostable β-glucosidase gene from Thermoanaerobacter brockii exhibited 0.5 U/ml of β-glucosidase activities. The signal peptide sequence of lytF gene from Bacillus subtilis was the most appropriate for the β-glucosidase secretion from Brevibacillus choshinensis, and the broth exhibited 0.74 U/ml of β-glucosidase activities. The engineered E. coli and B. choshinensis expressing the thermostable β-glucosidase gene produced 47.4 g/L glucose and 49.4 g/L glucose, respectively. Glucose was produced by the hydrolysis of 100 g/L Avicel cellulose for 10 days through BSES, and the product yield was similar to that obtained through BSES with purified β-glucosidase supplementation. Our findings indicate that the direct supplementation of β-glucosidase using bacterial cells expressing β-glucosidase gene or their broth was applicable to BSES, suggesting the potential of this process as a cost-effective approach to cellulose saccharification.

Keywords: Biological simultaneous enzyme production and saccharification; Biorefinery; Cellulosic biomass; Clostridium thermocellum; β-Glucosidase.