Acinetobacter baylyi ADP1 as a Platform for Flavonoid Production: Optimising homoeriodictyol and naringenin production
Mäkelä, Aarni (2025)
2025
Bioteknologian ja biolääketieteen tekniikan maisteriohjelma - Master's Programme in Biotechnology and Biomedical Engineering
Lääketieteen ja terveysteknologian tiedekunta - Faculty of Medicine and Health Technology
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Hyväksymispäivämäärä
2025-12-23
Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:tuni-2025122212078
https://urn.fi/URN:NBN:fi:tuni-2025122212078
Tiivistelmä
Flavonoids offer several beneficial properties for medical and industrial applications. However, their production has primarily relied on chemical extraction from plants, which poses challenges such as low yield and the use of harmful chemicals. Producing flavonoids using genetically engineered microbes could address these issues in industrial production. Microbes exhibit rapid production cycles, and engineered cell factories enable highly specific synthesis under chemically mild conditions without requiring extensive land use or dependence on climate conditions.
The bacterium Acinetobacter baylyi ADP1 is a promising host organism for flavonoid production, as it can utilise lignin-derived aromatic compounds as a carbon source. Lignin is a highly abundant and renewable resource that provides precursors required for flavonoid biosynthesis, such as p-coumaric acid and ferulic acid. Thus, A. baylyi ADP1 could be used to valorise lignin-derived aromatic compounds into flavonoids, enabling more environmentally sustainable production. Moreover, the straightforward genetic engineering of this strain makes it ideal for synthetic biology research.
In this thesis, A. baylyi ADP1 is employed as a production host for two flavonoids with promising pharmaceutical properties: naringenin and homoeriodictyol. The objectives were to improve naringenin production in this organism and to enable homoeriodictyol synthesis from ferulic acid. The study applies metabolic engineering techniques and constructs an A. baylyi ADP1 strain containing an autonomous genetic switch for dynamic regulation of flavonoid production. Additionally, the effects of four different mutant versions of chalcone synthase (CHS), a key enzyme in the flavonoid biosynthesis pathway, are investigated for improved naringenin production. The biosynthetic pathway for homoeriodictyol production is established by constructing two production strains using genes from three plant species: Hordeum vulgare, Stelonoma chusanum, and Medicago sativa. The engineered bacterial strains are evaluated for flavonoid production under various cultivation conditions, including fed-batch bioreactor experiments.
The strains were successfully constructed according to the objectives. To the best of my knowledge, this is the first report of homoeriodictyol production in A. baylyi ADP1. Both constructed strains produced homoeriodictyol from ferulic acid, with the highest product titre, 2.5 mg/L, achieved in batch culture at 20 °C. Furthermore, all tested CHS mutant strains produced naringenin, although yields were lower compared to the original enzyme. While the implemented dynamic genetic switch did not yield the hypothesised outcome, it offers interesting targets for further research to optimise flavonoid production. This thesis takes the first steps toward producing multiple flavonoids in A. baylyi ADP1 and lays the foundation for using lignin-derived aromatic compounds for the environmentally sustainable production of these valuable, healthpromoting compounds.
The bacterium Acinetobacter baylyi ADP1 is a promising host organism for flavonoid production, as it can utilise lignin-derived aromatic compounds as a carbon source. Lignin is a highly abundant and renewable resource that provides precursors required for flavonoid biosynthesis, such as p-coumaric acid and ferulic acid. Thus, A. baylyi ADP1 could be used to valorise lignin-derived aromatic compounds into flavonoids, enabling more environmentally sustainable production. Moreover, the straightforward genetic engineering of this strain makes it ideal for synthetic biology research.
In this thesis, A. baylyi ADP1 is employed as a production host for two flavonoids with promising pharmaceutical properties: naringenin and homoeriodictyol. The objectives were to improve naringenin production in this organism and to enable homoeriodictyol synthesis from ferulic acid. The study applies metabolic engineering techniques and constructs an A. baylyi ADP1 strain containing an autonomous genetic switch for dynamic regulation of flavonoid production. Additionally, the effects of four different mutant versions of chalcone synthase (CHS), a key enzyme in the flavonoid biosynthesis pathway, are investigated for improved naringenin production. The biosynthetic pathway for homoeriodictyol production is established by constructing two production strains using genes from three plant species: Hordeum vulgare, Stelonoma chusanum, and Medicago sativa. The engineered bacterial strains are evaluated for flavonoid production under various cultivation conditions, including fed-batch bioreactor experiments.
The strains were successfully constructed according to the objectives. To the best of my knowledge, this is the first report of homoeriodictyol production in A. baylyi ADP1. Both constructed strains produced homoeriodictyol from ferulic acid, with the highest product titre, 2.5 mg/L, achieved in batch culture at 20 °C. Furthermore, all tested CHS mutant strains produced naringenin, although yields were lower compared to the original enzyme. While the implemented dynamic genetic switch did not yield the hypothesised outcome, it offers interesting targets for further research to optimise flavonoid production. This thesis takes the first steps toward producing multiple flavonoids in A. baylyi ADP1 and lays the foundation for using lignin-derived aromatic compounds for the environmentally sustainable production of these valuable, healthpromoting compounds.