Adaptive laboratory evolution of <i>Clostridium autoethanogenum</i> to metabolize CO <sub>2</sub> and H <sub>2</sub> enhances growth rates in chemostat and unravels proteome and metabolome alterations
James K. Heffernan, R. Axayactl Garcia Gonzalez, Vishnu Mahamkali, Tim McCubbin, Venea Dara Daygon, Lian Liu, Robin Palfreyman, Audrey Harris, Michael Koepke, Kaspar Valgepea, Lars K. Nielsen, Esteban Marcellin
Abstract
Abstract Gas fermentation of CO 2 and H 2 is an attractive means to sustainably produce fuels and chemicals. Clostridium autoethanogenum is a model organism for industrial CO to ethanol and presents an opportunity for CO 2 ‐to‐ethanol processes. As we have previously characterized its CO 2 /H 2 chemostat growth, here we use adaptive laboratory evolution (ALE) with the aim of improving growth with CO 2 /H 2 . Seven ALE lineages were generated, all with improved specific growth rates. ALE conducted in the presence of 2% CO along with CO 2 /H 2 generated Evolved lineage D, which showed the highest ethanol titres amongst all the ALE lineages during the fermentation of CO 2 /H 2 . Chemostat comparison against the parental strain shows no change in acetate or ethanol production, while Evolved D could achieve a higher maximum dilution rate. Multi‐omics analyses at steady state revealed that Evolved D has widespread proteome and intracellular metabolome changes. However, the uptake and production rates and titres remain unaltered until investigating their maximum dilution rate. Yet, we provide numerous insights into CO 2 /H 2 metabolism via these multi‐omics data and link these results to mutations, suggesting novel targets for metabolic engineering in this bacterium.