Examinando por Materia "Cupriavidus necator"

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    Evaluation of metabolic capabilities of Cupriavidus necator to produce polyhydroxyalkanoates using low-cost carbon sources
    (Universidad del Valle, 2026) Ascencio Galván, Martha Ligia; Ramirez-Malule, Howard; López Agudelo, Víctor Alonso; Gómez Ríos, David
    Cupriavidus necator (C. necator) is a bacterium of great biotechnological interest due to its ability to synthesize polyhydroxyalkanoates (PHA), which are biodegradable polymers with industrial applications. The main objective of this study was to evaluate the metabolic capabilities of C. necator for PHA production by integrating different experimental conditions and computational analyses. To achieve this, batch and pulse-fed batch cultures were carried out. For the latter, three different feeding strategies were applied. In the first strategy, two pulses (8.33 mL) of dextrose derived from bitter cassava starch (20 g/L) and ammonium sulfate at a C/N ratio of 73 were added at 24 and 48 h, extending the culture to 120 h. In the second strategy, a single pulse (3.15 mL) of only dextrose (20 g/L) was added at 48 h, with a total cultivation time of 96 h. And in the third strategy, a single pulse (6.14 mL) of only dextrose (20 g/L) was added at 60 h, also extending the culture to 96 h. Batch cultures demonstrated the growth of C. necator, reaching a maximum biomass concentration of 5.17 g/L at 48 h and a polyhydroxybutyrate (PHB) accumulation of 47.1% at the end of the culture (72 h). In the pulse-fed batch cultures, the first strategy extended the exponential growth phase. This led to a gradual accumulation of PHB, reaching a PHB yield of 0.46 g/g and a productivity of 15 mg/(L*h) after 120 h of cultivation. However, the pulse-fed batch cultures, the first strategy resulted in an extended exponential growth phase and gradual PHB accumulation, reaching yields of 0.46 g/g and a productivity of 15.00 mg/(L*h) after 120 hours of cultivation. However, PHB accumulation was 20.57% and 25.12% lower compared to the accumulation observed in the batch culture. In contrast, the third strategy, with a single pulse at 60 h during the stationary phase, resulted in a higher PHB accumulation (52.40%) and productivity (23.22 mg/(L*h)), surpassing the values obtained in batch and the other pulse-feeding strategies. Additionally, the iCNH2025A model represents a refined and highly curated metabolic reconstruction of C. necator, showing significant improvements compared to previous models. With 2,737 reactions, 1,803 unique metabolites, and 1,059 annotated genes, the model integrates key metabolic pathways, including the biosynthesis of PHB and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), as well as central metabolic routes such as the Calvin-Benson-Bassham cycle (CBB), the Entner-Doudoroff (ED) pathway, the Embden-Meyerhof-Parnas (EMP) pathway, and the non-oxidative branch of the pentose phosphate pathway (PPP). The model was validated using MEMOTE, achieving a score of 89% and outperforming previous reconstructions such as RehMBEL1391 and iCN1361. In silico growth simulations with fructose and dextrose showed a high correlation with experimental data (R² = 0.76–0.78), validating the model's predictive capacity. Moreover, it accurately captured the carbon flux toward PHB synthesis under various experimental conditions, consistent with previous studies. Thus, the iCNH2025A model represents a significant advancement in the metabolic modeling of C. necator, offering a highly accurate and validated tool for analyzing its metabolism. Its ability to predict metabolic fluxes and growth rates makes it a valuable resource for industrial biotechnology, supporting future optimization strategies in biopolymer production.