Understanding Clostridium acetobutylicum ATCC 824 Metabolism Using Genome-Scale Thermodynamics and Metabolomics-based Modeling

Biobutanol has a potential application as a biofuel and can replace gasoline as an alternative fuel. However, the productivity of the biobutanol process has to be increased significantly, before it can be commercialized. Clostridium acetobutylicum has been the primary microbial host used for biobutanol production. Consequently, systems biology based genome-scale metabolic model of C. acetobutylicum metabolism is essential to optimize the biobutanol production rate via genetic and process optimization and metabolic engineering. An updated genome-scale metabolic model of C. acetobutylicum ATCC824 consists of 700 genes, 709 reactions, 29 exchange reactions, and 679 metabolites, which covers 16.3% of ORFs and is more comprehensive compared to two other existing models. This metabolic network was used as a platform for simulating phenotypes using the constraint-based modeling approach. Flux variability analysis shows the presence of alternate carbon and electron sinks that allows for different carbon assimilation patterns and ranges of product formation rates. Incorporation of metabolomics and thermodynamics data into this model results in thermodynamically feasible flux distributions. We apply an NMR based approach to quantify 31 intracellular metabolites in both metabolic phases. The total molar concentration of metabolites was approximately 245mM, where asparagine was the most dominant metabolite.