Structural Metabolic Modelling of the Acetogen Acetobacterium woodii

Acetogens are microbes that produce acetate as a fermentation by-product from a diverse range of substrates including gaseous substrates. They achieve this by employing the Wood- Ljungdahl Pathway (WLP), which enables the xation of carbon dioxide into cellular material for growth and to generate energy via a non-photosynthetic route. Electrons for this process are derived from diverse substrates that include molecular hydrogen and carbon monoxide. The ability of acetogens to utilise synthesis gas (H2, CO, CO2) make them an attractive target for industrial biotechnology since these are potentially renewable feedstocks. A model acetogen is Acetobacterium woodii, but as of yet no computational analysis has been carried out on it. Here we construct metabolic models, including a Genome-scale Metabolic Model (GSM) of Acetobacterium woodii using the sequenced and annotated genome of strain DSM1030. The GSM consists of 867 metabolites, 841 reactions and 98 transporters and is predictive of growth from the diverse substrates utilised by A. woodii . It is validated against experimental data produced here and that reported in the scientic literature. To analyse the impact of producing heterologous compounds with A. woodii , a core model was constructed from the GSM and then used in conjunction with heterologous synthesis pathways for elementary modes analysis. The GSM was also used to analyse how growth would be affected by heterologous compound production. For this effort, a combination of linear programs were written that consistently calculated a Ymax/ATP. It was found that no single LP could consistently calculate a Ymax/ATP. ATP calculations were also used to compare A. woodii maintenance costs across several conditions. Elementary modes analysis was carried out on a smaller WLP-only model, the results of which were used to interpret seemingly inconsistent reports on CO-dependent growth of A. woodii . Additionally it is highlighted that an as of yet unknown regulatory framework is likely biasing acetate production in conditions that could otherwise produce other compounds. Novel biosynthesis routes for commodity chemicals are also reported here and analysed for their impact on A. woodii metabolism along with other synthetic routes reported in the scientic literature and patents. Based on modelling results, suggestions are made for metabolically engineering A. woodii for applications in industrial biotechnology