A genome-scale metabolic network of the aroma bacterium Leuconostoc mesenteroides subsp. cremoris
Leuconostoc mesenteroides subsp. cremoris is an obligate heterolactic fermentative lactic acid bacterium that is mostly used in industrial dairy fermentations. The phosphoketolase pathway (PKP) is a unique feature of the obligate heterolactic fermentation, which leads to the production of lactate, ethanol, and/or acetate, and the final product profile of PKP highly depends on the energetics and redox state of the organism. Another characteristic of the L. mesenteroides subsp. cremoris is the production of aroma compounds in dairy fermentation, such as in cheese production, through the utilization of citrate. Considering its importance in dairy fermentation, a detailed metabolic characterization of the organism is necessary for its more efficient use in the industry. To this aim, a genome-scale metabolic model of dairy-origin L. mesenteroides subsp. cremoris ATCC 19254 (iLM.c559) was reconstructed to explain the energetics and redox state mechanisms of the organism in full detail. The model includes 559 genes governing 1088 reactions between 1129 metabolites, and the reactions cover citrate utilization and citrate-related flavor metabolism. The model was validated by simulating co-metabolism of glucose and citrate and comparing the in silico results to our experimental results. Model simulations further showed that, in co-metabolism of citrate and glucose, no flavor compounds were produced when citrate could stimulate the formation of biomass. Significant amounts of flavor metabolites (e.g., diacetyl and acetoin) were only produced when citrate could not enhance growth, which suggests that flavor formation only occurs under carbon and ATP excess. The effects of aerobic conditions and different carbon sources on product profiles and growth were also investigated using the reconstructed model. The analyses provided further insights for the growth stimulation and flavor formation mechanisms of the organism.
KeywordsLactic acid bacteria Leuconostoc mesenteroides subsp. cremoris Heterolactic fermentation Flavor metabolism Genome-scale metabolic model Flux balance analysis
Burcu Şirin (Yeditepe University) and Sebastián N. Mendoza (VU Amsterdam) are acknowledged for their help in HPLC analyses and reorganizing the SBML file, respectively.
This work received financial support from The Scientific and Technological Research Council of Turkey through TUBITAK 2214-A program and the Marmara University Scientific Research Project Fund through Project No: FEN-C-DRP-091116-0498.
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Conflict of interest
The authors declare that they have no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
- Chun BH, Kim KH, Jeon HH, Lee SH, Jeon CO (2017) Pan-genomic and transcriptomic analyses of Leuconostoc mesenteroides provide insights into its genomic and metabolic features and roles in kimchi fermentation. Sci Rep 7(1):11504. https://doi.org/10.1038/s41598-017-12,016-z CrossRefPubMedPubMedCentralGoogle Scholar
- Flahaut NAL, Wiersma A, van de Bunt B, Martens DE, Schaap PJ, Sijtsma L, dos Santos VAM, de Vos WM (2013) Genome-scale metabolic model for Lactococcus lactis MG1363 and its application to the analysis of flavor formation. Appl Microbiol Biotechnol 97(19):8729–8739. https://doi.org/10.1007/s00253-013-5140-2 CrossRefPubMedGoogle Scholar
- Frantzen CA, Kot W, Pedersen TB, Ardo YM, Broadbent JR, Neve H, Hansen LH, Dal Bello F, Ostlie HM, Kleppen HP, Vogensen FK, Holo H (2017) Genomic characterization of dairy associated Leuconostoc species and diversity of Leuconostocs in undefined mixed mesophilic starter cultures. Front Microbiol 8:132. https://doi.org/10.3389/fmicb.2017.00132 CrossRefPubMedPubMedCentralGoogle Scholar
- Garvie EI (1986) Genus Leuconostoc. In: Sneath PHA, Mair NS, Sharpe ME, Holt JG (eds) Bergey’s manual of systematic bacteriology, 2nd edn. Springer-Verlag, New YorkGoogle Scholar
- Koduru L, Kim Y, Bang J, Lakshmanan M, Han NS, Lee DY (2017) Genome-scale modeling and transcriptome analysis of Leuconostoc mesenteroides unravel the redox governed metabolic states in obligate heterofermentative lactic acid bacteria. Sci Rep 7(1):15721. https://doi.org/10.1038/s41598-017-16026-9 CrossRefPubMedPubMedCentralGoogle Scholar
- Naylor CE, Gover S, Basak AK, Cosgrove MS, Levy HR, Adams MJ (2001) NADP+ and NAD+ binding to the dual coenzyme specific enzyme Leuconostoc mesenteroides glucose 6-phosphate dehydrogenase: different interdomain hinge angles are seen in different binary and ternary complexes. Acta Crystallogr D Biol Crystallogr 57(Pt 5):635–648CrossRefGoogle Scholar
- Nielsen J (2017) Systems biology of metabolism. Annu Rev Biochem 86:245–275. https://doi.org/10.1146/annurev-biochem-061516-044757 CrossRefPubMedGoogle Scholar
- Pedersen MB, Gaudu P, Lechardeur D, Petit MA, Gruss A (2012) Aerobic respiration metabolism in lactic acid bacteria and uses in biotechnology. Annu Rev Food Sci Technol 3:37–58. https://doi.org/10.1146/annurev-food-022811-101,255 CrossRefPubMedGoogle Scholar
- Schellenberger J, Que R, Fleming RM, Thiele I, Orth JD, Feist AM, Zielinski DC, Bordbar A, Lewis NE, Rahmanian S, Kang J, Hyduke DR, Palsson BO (2011) Quantitative prediction of cellular metabolism with constraint-based models: the COBRA Toolbox v2.0. Nat Protoc 6(9):1290–1307. https://doi.org/10.1038/nprot.2011.308 CrossRefPubMedPubMedCentralGoogle Scholar
- Teusink B, van Enckevort FH, Francke C, Wiersma A, Wegkamp A, Smid EJ, Siezen RJ (2005) In silico reconstruction of the metabolic pathways of Lactobacillus plantarum: comparing predictions of nutrient requirements with those from growth experiments. Appl Environ Microbiol 71(11):7253–7262. https://doi.org/10.1128/AEM.71.11.7253-7262.2005 CrossRefPubMedPubMedCentralGoogle Scholar
- Tracey RP, Britz TJ (1989) Cellular fatty-acid composition of Leuconostoc oenos. J Appl Bacteriol 66(5):445–456. https://doi.org/10.1111/j.1365-2672.1989.tb05114.x CrossRefGoogle Scholar