A metabolic reconstruction of Lactobacillus reuteri JCM 1112 and analysis of its potential as a cell factory

• Published in: bioRxiv
• Main Authors: Kristjansdottir, Thordis, Bosma, Elleke F, Santos, Filipe, Özdemir, Emre, Herrgård, Markus J, França, Lucas, Bruno Sommer Ferreira, Nielsen, Alex T, Guđmundsson, Steinn
• Format: Paper
• Language: English
• Published: Cold Spring Harbor Cold Spring Harbor Laboratory Press 01.08.2019
• Subjects: 1,3-Propanediol; 3-Hydroxypropionaldehyde; Ethanol; Fermentation; Fermented food; Genomes; Glycerol; Glycolysis; Growth rate; Lactic acid; Lactobacillus reuteri; Metabolic engineering; Metabolism; Metabolites; Phosphoketolase; Probiotics; Secretion
• DOI: 10.1101/708875

Background: Lactobacillus 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. Results: A 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. Conclusion: We 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. Footnotes * https://github.com/steinng/reuteri/