Pathway engineering results the altered polyhydroxyalkanoates composition in recombinant Escherichia coli

Area of Science:

• Biotechnology
• Metabolic Engineering
• Polymer Science

Background:

• Polyhydroxyalkanoates (PHAs) are biodegradable polyesters with diverse applications.
• Efficient production of SCL-MCL PHAs from mixed substrates remains a challenge.
• Metabolic engineering of Escherichia coli offers a promising platform for PHA synthesis.

Purpose of the Study:

• To engineer a metabolically modified Escherichia coli strain for efficient production of SCL-MCL PHAs copolymers.
• To investigate the utilization of mixed carbon sources (sugars and fatty acids) for PHA synthesis.
• To analyze the impact of genetic modifications and substrate composition on PHA monomer composition.

Main Methods:

• Construction of a fadA gene mutant in a phosphotransferase system (PTS) disrupted E. coli DH5α strain.
• Transformation of the engineered E. coli with plasmids expressing PHA synthase genes (pCJY02, pBHR68, pBHR71).
• Cultivation of recombinant strains in media containing glucose and/or decanoate, followed by PHA analysis.

Main Results:

• The pCJY02-harboring recombinant accumulated SCL-MCL PHAs (3HB, 3HHx, 3HO, 3HD) from decanoate with 1.90 wt% copolymer content.
• When utilizing a decanoate and glucose mixture, the recombinant produced SCL-MCL PHAs with significantly higher content (7.3 wt%) and altered monomer ratios.
• The PHA synthase PhaC2(Ps) enabled the synthesis of PHA copolymers from mixed substrates, with composition modulated by carbon source availability.

Conclusions:

• Engineered E. coli DH5α (ΔptsG, ΔFadA) with pCJY02 can efficiently produce SCL-MCL PHAs copolymers from mixed sugars and fatty acids.
• The low-substrate-specificity PHA synthase PhaC2(Ps) is crucial for synthesizing PHA copolymers from diverse carbon sources.
• Metabolic engineering strategies and control of carbon source composition allow for modulation of PHA monomer composition and properties.