Fig 1.
Flowchart of the dynamic flux balance analysis (dFBA) framework used to describe the metabolic dynamics of Streptomyces clavuligerus growing in GLYCAS-5 medium.
The scheme summarizes experimental inputs, model constraints, and simulation outputs integrated in the modeling workflow.
Fig 2.
Amino acid composition of the GLYCAS-5 medium.
Quantitative analysis of amino acid concentrations (mg/L) determined from three independent biological replicates (n = 3). Values are reported as mean concentrations.
Fig 3.
Growth kinetics of Streptomyces clavuligerus ATCC 27064 in the GLYCAS-5 medium.
Concentrations were determined from three independent biological replicates (n = 3). Growth phases are indicated as follows: EEP, early exponential phase; EP, exponential phase (grey zone); SP, stationary phase; DP, death phase; TP, trophophase; IP, idiophase.
Fig 4.
Dynamics of amino acid consumption by Streptomyces clavuligerus ATCC 27064 during growth in GLYCAS-5 medium.
Concentrations were determined from three independent biological replicates (n = 3). (A) Glutamate (L-Glu), Aspartate (L-Asp), Proline (L-Pro), and Lysine (L-Lys); (B) Leucine (L-Leu), Serine (L-Ser), Phenylalanine (L-Phe), and Alanine (L-Ala); (C) Threonine (L-Thr), Cysteine (L-Cys), Arginine (L-Arg), and Glycine (L-Gly); (D) Tyrosine (L-Tyr), Valine (L-Val), Histidine (L-His), and Tryptophan (L-Trp).
Table 1.
Estimated kinetic parameters for the unstructured dynamic model describing Streptomyces clavuligerus growth and CA production in GLYCAS-5 medium.
Fig 5.
Topological analysis of the amino acid metabolic network derived from the iLT1021 GEM.
(A) Central hub metabolites and their interaction map, (B) Correlation heatmap of network topology parameters, (C) Quantitative distribution of connectivity metrics across amino acids.
Fig 6.
Nitrogen source effect on clavulanic acid biosynthesis and growth using glycerol as a sole carbon source (1 mmol/gDW·h).
The intersection of different growth profiles, on specific amino acids, corresponds to the best scenario for CA synthesis while maintaining a minimum Streptomyces clavuligerus biomass growth.
Fig 7.
Dynamic flux balance analysis (dFBA) of S. clavuligerus in GLYCAS-5 medium.
(A) Metabolic fluxes during early exponential (yellow), stationary (red), and decay (blue) phases. Phases are demarcated by glycerol depletion and a decrease in growth rate. (B) Oxygen uptake and CO₂ secretion fluxes. (C) CA secretion flux. (D) Carbon flux distribution contributing to CA biosynthesis in decay phase (DP).
Fig 8.
Temporal metabolic flux profiles in Streptomyces clavuligerus during dFBA simulation.
Dynamics of amino acid metabolism (A–D), glycolytic pathway (E–K), tricarboxylic acid (TCA) cycle (L–M), pentose phosphate pathway (N–O), nitrogen metabolism (P–R), and precursor metabolism for secondary metabolites. Early exponential (yellow), stationary (red), and decay (blue) phases are indicated. Enzymes: Transaldolase (TALA), transketolase (TKT1/2), glucose-6-phosphate isomerase (PGI), triose phosphate isomerase (TPI), ribose-5-phosphate isomerase (RPI), ribulose-5-phosphate epimerase (RPE), fructose-1,6-bisphosphate (FBP), aconitase (ACONT), isocitrate dehydrogenase (ICDH), fumarase (FUM), malic enzyme (ME), succinate dehydrogenase (SUCD1), malate dehydrogenase (MDH-2), glutamine synthetase (GS), glutamate dehydrogenase (GDH), glutamate oxaloacetate transaminase (GOT), carbamoyl phosphate synthetase (CPSase), and argininosuccinate synthetase (ARGSS).