(191ad) System Metabolic Engineering of Clostridium Cellulovorans Towards Consolidated Bioprocessing for N-Butanol Production from Cellulosic Biomass

n-Butanol can be used as a significant industrial chemical and potential superior biofuel. Especially, compared with ethanol, n-butanol can be used as an ideal substitute of gasoline because of its high energy density, low water solubility, and low vapor pressure. Lignocellulosic biomass is an abundant, cheap, and renewable carbon source, and is a desirable feedstock for biofuels production. In our previous research, Clostridium cellulovorans, a natural cellulose/hemicelluloses utilizing, butyrate producing bacterium, was metabolically engineered for n-butanol production from cellulose by overexpressing adhE2 from Clostridium acetobutylicum. Therefore, consolidated bioprocessing (CBP) with combined enzyme production, biomass hydrolysis, and sugar fermentation in one step can be achieved by using the engineered C. cellulovorans. However, this mutant only produced 1.42 g/L n-butanol and 1.60 g/L ethanol from microcrystalline cellulose in 10 days. In this study, genome-scale metabolic model with omics analysis was firstly applied for guiding rational metabolic engineering of C. cellulovorans. System metabolic and process engineering, including optimization of butanol biosynthesis pathway, increasing C4 carbon flux, weakening competitive acids synthesis pathway, and improving the intracellular NADH availability through redox rebalancing, were carried out to increase butanol production in C. cellulovorans. The mutant produced 5.2 g/L n-butanol with a high yield of >0.3 g/g cellulose in batch fermentation, which were the highest ever achieved. This study demonstrated that C. cellulovorans is a promising CBP host for biobutanol production from cellulosic biomass.