Genome-scale metabolic modelling enables deciphering ethanol metabolism via the acrylatepathway in the propionate-producer Anaerotignum neopropionicum

Background: Microbial production of propionate from diluted streams of ethanol (e.g., deriving from syngas fermentation) is a sustainable alternative to the petrochemical production route. Yet, few ethanol-fermenting propionigenic bacteria are known, and understanding of their metabolism is limited. Anaerotignum neopropionicum is a propionate-producing bacterium that uses the acrylate pathway to ferment ethanol and CO2 to propionate and acetate. In this work, we used computational and experimental methods to study the metabolism of A. neopropionicum and, in particular, the pathway for conversion of ethanol into propionate. Results: Our work describes iANEO_SB607, the first genome-scale metabolic model (GEM) of A. neopropionicum. The model was built combining the use of automatic tools with an extensive manual curation process, and it was validated with experimental data from this and published studies. The model predicted growth of A. neopropionicum on ethanol, lactate, sugars and amino acids, matching observed phenotypes. In addition, the model was used to implement a dynamic flux balance analysis (dFBA) approach that accurately predicted the fermentation profile of A. neopropionicum during batch growth on ethanol. A systematic analysis of the metabolism of A. neopropionicum combined with model simulations shed light into the mechanism of ethanol fermentation via the acrylate pathway, and revealed the presence of the electron-transferring complexes NADH-dependent reduced ferredoxin:NADP+ oxidoreductase (Nfn) and acryloyl-CoA reductase-EtfAB, identified for the first time in this bacterium. Conclusions: The realisation of the GEM iANEO_SB607 is a stepping stone towards the understanding of the metabolism of the propionate-producer A. neopropionicum. With it, we have gained insight into the functioning of the acrylate pathway and energetic aspects of the cell, with focus on the fermentation of ethanol. Overall, this study provides a basis to further exploit the potential of propionigenic bacteria as microbial cell factories.