Presentation Information
[1Chemi-12]Carbon dioxide based 3-hydroxypropionic acid production
○Kang Lan Tee1 (1. University of Sheffield (UK))
Keywords:
Carbon dioxide,3-hydroxypropionic acid,Cupriavidus necator,Sustainable biomanufacturing,Engineering biology
The global market for the platform chemical 3-hydroxypropionic acid (3-HP) is rapidly expanding and is projected to reach USD 1.3 billion by 2030. This is driven by its diverse applications, especially as a sustainable precursor for bio-based acrylic acid. Traditionally, microbial production of 3-HP production has relied on glycerol-based or glucose-based pathways. In this study, we extended the substrate repertoire by engineering a CO2-based 3-HP producing strain of Cupriavidus necator.
To enable in vivo product accumulation, we knocked out native 3-HP catabolic genes in C. necator and introduced four distinct malonyl-CoA reductase (MCR) systems individually to establish a malonyl-CoA-dependent biosynthetic route. The enzyme systems investigated included the well-characterised bifunctional MCR from a green non-sulfur bacterium, an archaeal MCR system, a novel bifunctional bacterial MCR, and a synthetic ancestral MCR reconstructed through evolutionary design. Leveraging C. necator's substrate versatility, the engineered strains successfully synthesised 3-HP using sodium gluconate, glycerol and CO2 as carbon sources. All four MCR systems supported 3-HP synthesis, confirmed by HPLC analysis of the spent medium. The highest titre observed was 1.3 g/L in fed-batch fermentation using sodium gluconate. Interestingly, the ancestral MCR outperformed its natural homologue, and autotrophic 3-HP production from CO2 was demonstrated using gas fermentation.
These results reveal that,
1. multiple, evolutionarily diverse MCR systems can power efficient 3-HP biosynthesis;
2. synthetic ancestral enzyme can enhance productivity;
3. C. necator can directly convert CO2 into 3-HP, establishing a prospective carbon-negative route to sustainable acrylic acid.
To enable in vivo product accumulation, we knocked out native 3-HP catabolic genes in C. necator and introduced four distinct malonyl-CoA reductase (MCR) systems individually to establish a malonyl-CoA-dependent biosynthetic route. The enzyme systems investigated included the well-characterised bifunctional MCR from a green non-sulfur bacterium, an archaeal MCR system, a novel bifunctional bacterial MCR, and a synthetic ancestral MCR reconstructed through evolutionary design. Leveraging C. necator's substrate versatility, the engineered strains successfully synthesised 3-HP using sodium gluconate, glycerol and CO2 as carbon sources. All four MCR systems supported 3-HP synthesis, confirmed by HPLC analysis of the spent medium. The highest titre observed was 1.3 g/L in fed-batch fermentation using sodium gluconate. Interestingly, the ancestral MCR outperformed its natural homologue, and autotrophic 3-HP production from CO2 was demonstrated using gas fermentation.
These results reveal that,
1. multiple, evolutionarily diverse MCR systems can power efficient 3-HP biosynthesis;
2. synthetic ancestral enzyme can enhance productivity;
3. C. necator can directly convert CO2 into 3-HP, establishing a prospective carbon-negative route to sustainable acrylic acid.
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