Genome-scale metabolic model analysis of Pichia pastoris for enhancing the production of S-adenosyl-L-methionine.

Komagataella phaffii, formerly Pichia pastoris (P. pastoris), is a promising methylotrophic yeast used in industry to produce recombinant protein and valuable metabolites. In this study, a genome-scale metabolic model (GEMs) was reconstructed and used to assess P. pastoris' metabolic capabilities for the production of S-adenosyl-L-methionine (AdoMet or SAM or SAMe) from individual carbon sources along with the addition of L-methionine. In a model-driven P. pastoris strain, the well-established genome-scale metabolic model iAUKM can be implemented to predict high valuable metabolite production. The model, iAUKM, was created by merging the previously published iMT1026 model and the draught model generated using Raven toolbox from the KEGG database which covered 2309 enzymatic reactions associated with 1033 metabolic genes and 1750 metabolites. The highly curated model was successful in capturing P. pastoris growth on various carbon sources, as well as AdoMet production under various growth conditions. Many overexpression gene targets for increasing AdoMet accumulation in the cell have been predicted for various carbon sources. Inorganic phosphatase (IPP) was one of the predicted overexpression targets as revealed from simulations using iAUKM. When IPP gene was integrated into P. pastoris, we found that AdoMet accumulation increased by 16% and 14% using glucose and glycerol as carbon sources, respectively. Our in silico results shed light on the factors limiting AdoMet production, as well as key pathways for rationalized engineering to increase AdoMet yield.