Engineering Pseudomonas chlororaphis HT66 for the Biosynthesis of Copolymers Containing 3‑Hydroxybutyrate and Medium-Chain-Length 3‑Hydroxyalkanoates

• Published in: Journal of agricultural and food chemistry Vol. 72; no. 15; pp. 8684 - 8692
• Main Authors: Deng, Ru-Xiang, Li, Hui-Ling, Wang, Wei, Hu, Hong-Bo, Zhang, Xue-Hong
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 17-04-2024
• Subjects: Biotechnology and Biological Transformations; β-oxidation; synthetic biology; Pseudomonas chlororaphis; metabolic engineering; polyhydroxyalkanoate
• ISSN: 0021-8561; 1520-5118
• DOI: 10.1021/acs.jafc.4c00777

Polyhydroxyalkanoates (PHAs) are promising alternatives to petroleum-based plastics, owing to their biodegradability and superior material properties. Here, the controllable biosynthesis of scl-co-mcl PHA containing 3-hydroxybutyrate (3HB) and mcl 3-hydroxyalkanoates was achieved in Pseudomonas chlororaphis HT66. First, key genes involved in fatty acid β-oxidation, the de novo fatty acid biosynthesis pathway, and the phaC1-phaZ-phaC2 operon were deleted to develop a chassis strain. Subsequently, an acetoacetyl-CoA reductase gene phaB and a PHA synthase gene phaC with broad substrate specificity were heterologously expressed for producing and polymerizing the 3HB monomer with mcl 3-hydroxyalkanoates under the assistance of native β-ketothiolase gene phaA. Furthermore, the monomer composition of scl-co-mcl PHA was regulated by adjusting the amount of glucose and dodecanoic acid supplemented. Notably, the cell dry weight and scl-co-mcl PHA content reached 14.2 g/L and 60.1 wt %, respectively, when the engineered strain HT11Δ::phaCB was cultured in King’s B medium containing 5 g/L glucose and 5 g/L dodecanoic acid. These results demonstrated that P. chlororaphis can be a platform for producing scl-co-mcl PHA and has the potential for industrial application.