Traditionally, metabolic models only predict metabolic fluxes. Although this is directly applicable to the metabolites that we want to produce, such as DMDS, this does not suffice to answer our questions regarding the impact of the whole system. Therefore, we added the production of all the proteins of interest as if they are metabolites. For this we used the protein sequences to determine the correct stoichiometric ratio of amino acids [18]. The other side-substrates/products are in the same ratio as described for production of proteins for the biomass reaction in the GSMM. In the model Promoter Design the protein production rate is estimated.
As our metabolites of interest (DAPG, DMDS, DMTS) were not yet included in the GSMM, we had to deduce their production pathways based on databases and literature information [19,20,21,22](pathways and metabolites). Furthermore, we took special care that all added pathways are stoichiometrically balanced, which is essential for proper and reliable integration in a GSMM. To get the most realistic results we need to set a biologically relevant production rate. This can be done by estimating a production range with in vivo data from literature.