Integrated, systems metabolic picture of acetone-butanol-ethanol fermentation by Clostridium acetobutylicum

Abstract

Microbial metabolism involves complex, system-level processes implemented via the orchestration of metabolic reactions, gene regulation, and environmental cues. One canonical example of such processes is acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum, during which cells convert carbon sources to organic acids that are later reassimilated to produce solvents as a strategy for cellular survival. The complexity and systems nature of the process have been largely underappreciated, rendering challenges in understanding and optimizing solvent production. Here, we present a system-level computational framework for ABE fermentation that combines metabolic reactions, gene regulation, and environmental cues. We developed the framework by decomposing the entire system into three modules, building each module separately, and then assembling them back into an integrated system. During the model construction, a bottom-up approach was used to link molecular events at the singlecell level into the events at the population level. The integrated model was able to successfully reproduce ABE fermentations of the WT C. acetobutylicum (ATCC 824), as well as its mutants, using data obtained from our own experiments and from literature. Furthermore, the model confers successful predictions of the fermentations with various network perturbations across metabolic, genetic, and environmental aspects. From foundation to applications, the framework advances our understanding of complex clostridial metabolism and physiology and also facilitates the development of systems engineering strategies for the production of advanced biofuels.integrated modeling | ABE fermentation | clostridial physiology | systems biology | metabolic engineering M icrobial metabolism is a means by which a microbe uses nutrients and generates energy to live and reproduce. As one of the most fundamental cellular characteristics, it typically involves complex biochemical processes implemented through the orchestration of metabolic reactions and gene regulation, as well as their interactions with environmental cues (1-3). One representative example of such complex processes is solvent production by Clostridium acetobutylicum, a Gram-positive, anaerobic bacterium that is considered to be one of the most prominent species for industrial biofuel production (4).Solvent [acetone-butanol-ethanol (ABE)] fermentation of the species involves two physiological phases (5-8): During the first phase, the bacterium grows exponentially, and organic acids (acetic acid and butyric acid) are produced with the release of energy-the acidogenic phase. This process causes a dramatic drop in extracellular pH. In response to the substantial decrease of the pH, cells enter the stationary phase, and the organic acids formed are reassimilated to produce solvents including acetone, butanol, and ethanol-the solventogenic phase-thereby helping the bacterium to relieve the stress as a strategy for survival. Solventogenesis is subsequently accompanied by the onset of sporulati...

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