Constraint-Based Metabolic Model Elucidates Energy, Reducing Power, and Carbon Flux Distribution in Clostridium tyrobutyricum for Optimization of n-Butanol Production

n-Butanol is a promising alternative fuel source due to its high energy content, compatibility with combustion engines, and blending ability. However, it is currently more expensive to produce biobutanol from fermentation than petroleum-based butanol, and thus the biobutanol fuel market price is higher. Because the fermentation process is more sustainable and environmentally friendly, it is important to reduce the production cost of biobutanol by increasing butanol selectivity and yield through metabolic engineering of fermentative bacteria. The goal of this work was to determine a strategy to optimize Clostridium tyrobutyricum for biobutanol production. A constraint-based model of the central carbon metabolism of C. tyrobutyricum was developed from the network stoichiometry and enzymatic equilibrium constraints, which formed a linear optimization problem that was solved using various data sets and objective functions. The model was validated using butanol fermentation data, and the simulation results were employed to predict the optimal distribution of energy, reducing power, and carbon. Major factors limiting butanol production were identified, and several strategies for maximizing butanol selectivity and yield were proposed.