Supplementation of recombinant cellulases with LPMOs and CDHs improves consolidated bioprocessing of cellulose.

ABSTRACT

The increased demand for energy has sparked a global search for renewable energy sources that could partly replace fossil fuel resources and help mitigate climate change. Cellulosic biomass is an ideal feedstock for renewable bioethanol production, but the process is not currently economically feasible due to the high cost of pretreatment and enzyme cocktails to release fermentable sugars. Lytic polysaccharide monooxygenases (LPMOs) and cellobiose dehydrogenases (CDHs) are auxiliary enzymes that can enhance cellulose hydrolysis. In this study, four LPMO and two CDH genes were subcloned and expressed in the Saccharomyces cerevisiae Y294 laboratory strain. SDS-PAGE analysis confirmed the extracellular production of the LPMOs and CDHs in the laboratory S. cerevisiae Y294 strain. A rudimentary cellulase cocktail (cellobiohydrolase 1 and 2, endoglucanase and ß-glucosidase) was expressed in the commercial CelluX™ 4 strain and extracellular production of the individual cellulases was confirmed by SDS-PAGE analysis. In vitro cooperation of the CDHs and LPMOs with the rudimentary cellulases produced by strain CelluX™ 4[F4-1] was demonstrated on Whatman filter paper. The significant levels of soluble sugars released from this crystalline cellulose substrate indicated that these auxiliary enzymes could be important components of the CBP yeast cellulolytic system.