Redesigning E. Coli Metabolism for Obligate Anaerobic Production of Biofuels
The biological conversion of lignocellulosic biomass into biofuels has emerged as a promising route to sustainable energy. Fermentation is the most efficient route to synthesize biofuels that are reduced relative to fermentative sugars. However, most pathways that synthesize advanced biofuels are not obligately anaerobic. Thus, if we could redesign cell metabolism to use advanced biofuel-producing pathways as heterologous, obligate anaerobic pathways, we would achieve higher product yields and make large-scale production feasible and cost-effective because supply and precise control of oxygen are not required. To demonstrate the approach, elementary mode analysis was applied to redesign efficient Escherichia coli cells optimized for biofuels production such as short, medium, and long chain length alcohols from fermentative sugars under anaerobic conditions. The analysis decomposed the E. coli metabolic network into unique and elementary pathways from which only the optimal routes for alcohol production were selected. The designed cells were constrained to operate according to these selected pathways through multiple gene deletion, addition, and over-expression. In addition, the cells were specifically designed to tightly couple both biomass and alcohol production, which represented a unique property exploited for molecular pathway evolution to improve alcohol production and tolerance. Here we present the design, construction, and characterization of some of designed cells, and evaluate their performances with model prediction.