Biochemical and transcriptomic characterization of the bacterium Cupriavidus necator growing on volatile fatty acid mixtures: towards digestate utilization for bioplastics production.

The escalating global plastic pollution crisis highlights the urgent need for sustainable, biodegradable alternatives to petroleum-based plastics. Polyhydroxyalkanoates are promising biopolymers due to their excellent biocompatibility and biodegradability. While poly(3-hydroxybutyrate) is a common but brittle polyhydroxyalkanoates, the copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate), which incorporates 3-hydroxyvalerate units, offers superior mechanical properties and greater versatility. This thesis investigates the metabolically versatile bacterium Cupriavidus necator and its capacity for polyhydroxyalkanoate biosynthesis, with a focus on strategies to produce high-value poly(3-hydroxybutyrate-co-3-hydroxyvalerate). A distinct fermentation approach was employed, using glucose as a baseline reference and volatile fatty acids as a sustainable alternative carbon source, to produce poly(3-hydroxybutyrate) and poly(3-hydroxybutyrate-co-3-hydroxyvalerate), respectively. Controlled fed-batch fermentations revealed distinct kinetics: glucose-fed cultures exhibited rapid exponential growth, reaching 2.8–3.0 grams per liter of biomass, whereas volatile fatty acid-fed fermentations showed slower growth, achieving approximately 2.3–2.5 grams per liter. The resulting biopolymers were characterized, confirming successful production of both poly(3-hydroxybutyrate) and poly(3-hydroxybutyrate-co-3-hydroxyvalerate). Transcriptomic analysis further revealed that utilization of volatile fatty acids involves broad metabolic adaptations, including activation of the tricarboxylic acid cycle, oxidative phosphorylation, and transport systems. Overall, this work demonstrates the feasibility of producing biodegradable plastics from low-cost feedstocks and establishes a scalable bioprocess framework. It supports the valorization of waste-derived substrates into high-value bioplastics and contributes to the development of a circular bioeconomy.