Fig 1.
Hyaluronic acid biosynthesis pathway in S. zooepidemicus.
This schematic illustrates the biosynthetic pathway of hyaluronic acid (HA) emphasizing the metabolic precursors and enzymatic steps. The pathway starts from glucose, which is metabolized to form two activated sugar precursors: UDP-glucuronic acid (UDP-GA) and UDP-N-acetylglucosamine (UDP-GlcNAc). These precursors are polymerized by the enzyme hyaluronan synthase, encoded by the hasA gene, to form HA. Arrows indicate the direction of enzymatic reactions. Key energy inputs and cofactors are labeled. This figure contextualizes the metabolic investment required for HA synthesis, highlighting the critical nodes for regulation and genetic manipulation. Abbreviations: ATP, adenosine triphosphate; NAD, nicotinamide adenine dinucleotide; UTP, uridine triphosphate.
Table 1.
An overview of features for S. zooepidemicus metabolic model.
Table 2.
Features of iZN522 in comparison with other Streptococci metabolic models.
Table 3.
Comparison of predicted and experimental growth rate.
Table 4.
Predicted essential amino acids using iZN522 and comparison with experimental data [57].
Table 5.
The prediction of organism’s ability to consume various carbon source using iZN522 and comparison with experimental data [58].
Fig 2.
Robustness analysis of hyaluronic acid production under varying oxygen conditions.
Graph showing the predicted effect of varying oxygen exchange flux on hyaluronic acid production rate in S. zooepidemicus culture media containing yeast extract and sucrose. The robustness curve demonstrates increasing HA flux with increasing oxygen availability, consistent with experimental observations. The x-axis represents oxygen exchange rate (mmol/gDCW/h), and the y-axis represents predicted HA synthesis flux (mmol/gDCW/h). The figure illustrates oxygen’s role as a critical environmental factor enhancing HA biosynthesis in aerobic conditions.
Fig 3.
Results of in silico single-gene deletion analysis for iZN522 metabolic model.
Fig 4.
Identifying in silico multiple-mutant strain to improve HA production rate using iterative single-gene deletion analysis method: A: In silico gene deletions related to the asnA (encoding the asparagine synthetase) with the relative predicted HA production yield in moles of HA per mole of glucose B: In silico gene deletions related to asd (encoding the aspartate semi-aldehyde dehydrogenase) with relative predicted HA production yield in moles of HA per mole of glucose.
Table 6.
Predicted growth rate and HA production rate in single, double, and triple S. zooepidemicus in silico mutant strains.
Fig 5.
The metabolic flux distribution in ∆asd strain in comparison with the wild strain.
Black values indicate the reaction fluxes of the wild strain, and red values indicate reaction fluxes of ∆asd strain. The unit of fluxes is mmol.gDCW-1.h-1 The multiplication sign indicates the deletion of a gene. The equivalent abbreviations for important metabolites are given below (Udpg: UDP-glucose, Uacgam: UDP-N-acetyl glucosamine, PG: Peptidoglycan, Clpn-lla: Cardiolipin, Lyspg-lla: Lysyl phosphatidylglycerol, LTAalaGal: Lipoteichoic acid derivatives, HA: Hyaluronic acid).
Fig 6.
Comparison of metabolic flux distribution in ∆asd ∆ilvE strain with ∆asd strain.
Red values refer to the reaction fluxes of the ilvE ∆asd strain. The fluxes are reported in mmol.gCDW-1.h-1. (Udpg: UDP-glucose, Uacgam: UDP-N-acetylglucosamine, PG: peptidoglycan, Clpn-lla: cardiolipin, Lyspg-lla: lysyl phosphatidylglycerol, LTAalaGal: lipoteichoic acid derivatives, HA Hyaluronic acid, 3 mob: 3-methyl-2-oxobutanate, akg: alpha-ketoglutarate, glu: glutamate).
Fig 7.
Comparing the metabolic flux distribution in the ∆asd ∆ilvE ∆pyrD strain with the ∆asd ∆ilvE strain.
The black values represent the reaction fluxes of the ∆asd ∆ilvE strain. Red values refer to the reaction fluxes of the ∆asd ∆ilvE ∆pyrD strain. The fluxes are reported in mmol.gCDW-1.h-1. The cross indicates the deletion of the gene. (Udpg: UDP-glucose, Uacgam: UDP-N-acetyl glucosamine, PG: peptidoglycan, CPS-lla: polysaccharide units, Ura: uracil, Uri: uridine, GUA: guanine).
Fig 8.
Results of in silico single-gene deletion analysis for iCW773 metabolic model.
Table 7.
Results of single- and double-gene deletion analysis Streptococcus zooepidemicus and recombinant Corynebacterium glutamicum with flux balance analysis approach.
Fig 9.
Providing solutions to optimize the HA production rate based on multiple optimal solutions in the metabolic model of iCW773.
The blue arrows represent reactions that carry a normal flux; the green arrows should up-regulate, while the flux of reactions with the red arrows should down-regulate.
Fig 10.
The Comparison of flux distribution of central carbon metabolism pathway and HA production pathway in iZN522 and iCW773 metabolic networks.
S. zooepidemicus fluxes were reported in green, and recombinant C. glutamicum fluxes were shown in red. The dot line also indicates reactions that are present only in C. glutamicum, and S. zooepidemicus lacks them. Negative fluxes also indicate the reaction flux in the opposite direction.