Production of minicellulosomes from Clostridium cellulovorans for the fermentation of cellulosic ethanol using engineered recombinant Saccharomyces cerevisiae

Saccharomyces cerevisiae was engineered for assembly of minicellulosomes by heterologous expression of a recombinant scaffolding protein from Clostridium cellulovorans and a chimeric endoglucanase E from Clostridium thermocellum. The chimeric endoglucanase E fused with the dockerin domain of endoglucanase B from C. cellulovorans was assembled with the recombinant scaffolding protein. The resulting strain was able to ferment amorphous cellulose [carboxymethyl- cellulose (CMC)] into ethanol with the aid of β-glucosidase 1 produced from Saccharomycopsis fibuligera. The minicellulosomes assembled in vivo retained the synergistic effect for cellulose hydrolysis. The minicellulosomes containing the cellulose-binding domain were purified by crystalline cellulose affinity in a single step. In the fermentation test at 10 g L^-1 initial CMC, approximately 3.45 g L^-1 ethanol was produced after 16 h. The yield (in grams of ethanol produced per substrate) was 0.34 g g^-1 from CMC. This result indicates that a one-step processing of cellulosic biomass in a consolidated bioprocessing configuration is technically feasible by recombinant yeast cells expressing functional minicellulosomes.