A metabolic reconstruction ofLactobacillus reuteriJCM 1112 and analysis of its potential as a cell factory

Abstract

BackgroundLactobacillus reuteri is a heterofermentative Lactic Acid Bacterium (LAB) that is commonly used for food fermentations and probiotic purposes. Due to its robust properties, it is also increasingly considered for use as a cell factory. It produces several industrially important compounds such as 1,3-propanediol and reuterin natively, but for cell factory purposes, developing improved strategies for engineering and fermentation optimization is crucial. Genome-scale metabolic models can be highly beneficial in guiding rational metabolic engineering. Reconstructing a reliable and a quantitatively accurate metabolic model requires extensive manual curation and incorporation of experimental data.ResultsA genome-scale metabolic model of L. reuteri JCM 1112T was reconstructed and the resulting model, Lreuteri_530, was validated and tested with experimental data. Several knowledge gaps in the metabolism were identified and resolved during this process, including presence/absence of glycolytic genes. Flux distribution between the two glycolytic pathways, the phosphoketolase and Embden-Meyerhof-Parnas pathways, varies considerably between LAB species and strains. As these pathways result in different energy yields, it is important to include strain-specific utilization of these pathways in the model. We determined experimentally that the Embden-Meyerhof-Parnas pathway carried at most 7% of the total glycolytic flux. Predicted growth rates from Lreuteri_530 were in good agreement with experimentally determined values. To further validate the prediction accuracy of Lreuteri_530, the predicted effects of glycerol addition and adhE gene knock-out, which results in impaired ethanol production, were compared to in vivo data. Examination of both growth rates and uptake- and secretion rates of the main metabolites in central metabolism demonstrated that the model was able to accurately predict the experimentally observed effects. Lastly, the potential of L. reuteri as a cell factory was investigated, resulting in a number of general metabolic engineering strategies.ConclusionWe have constructed a manually curated genome-scale metabolic model of L. reuteri JCM 1112T that has been experimentally parameterized and validated and can accurately predict metabolic behavior of this important platform cell factory.

Recent biotechnological trends in lactic acid bacterial fermentation for food processing industries

“…Genome-scale metabolic models reconstruction can be beneficial for rational metabolic engineering. Kristjansdottir et al [177] reconstructed a genome model of L. reuteri JCM 1112 through flux distribution of glycolytic, phosphoketolase and Embden-Meyerhof-Parnas pathways. They performed extensive manual curation such as gap fillings, updating and gene-protein reaction associations, gene IDs update and updating reacting metabolites and reviewed the integration of organism-specific data.…”

Two-species community design of Lactic Acid Bacteria for optimal production of Lactate

“…and Pediococcus species were obtained (34). Previously curated and published GSMMs of L. plantarum WCSF1 and L. reuteri JCM 1112 were also used to construct the synthetic communities of LAB (14,30). A list of all 49 GSMMs used in this study is tabulated in Table S1.…”

“…L. pentosus consumed cellobiose, avoiding inhibition of T. reesei cellulase activity, and acetic acid produced from L. pentosus was utilised as a carbon source by the fungus (13). GSMMs of various LAB such as Lactobacillus reuteri, Leuconostoc mesenteroides, Lactobacillus plantarum, Lactobacillus casei, Lactococcus lactis, and Streptococcus thermophilus have been published (14).…”