Fig 1.
β-Alanine pathway used in this study.
Four L-aspartate decarboxylases (PanD) from E. coli, B. subtilis, P. fluorescens, and C. glutamicum were tested. PP0596, β-alanine-pyruvate transaminase from Pseudomonas putida; YdfG, 3-hydroxyacid dehydrogenase from E. coli; PrpE, propionyl-CoA synthase rom E. coli; PhaC1, polyhydroxyalkanoate synthase from Cupriavidus necator.
Table 1.
Bacterial strains, plasmids, and primers used in this study.
Fig 2.
Effect of β-alanine and L-aspartate on the P3HP production.
The strain Q2153 was grown in minimal medium with supplement of 5 g/L β-alanine (β-Ala) or L-aspartate (L-Asp), cultivation without amino acid was used as the control (CK), and the P3HP production (white), CDW (light grey), and P3HP content (heavy grey) were presented. The experiment was carried out in shaking flask in triplicate. All shake-flask experiments were incubated for 48 h after induction.
Fig 3.
Screening of L-aspartate decarboxylase for P3HP production.
P3HP-producing strains with different panD gene were grown in minimal medium, and the P3HP production (white), CDW (light grey), and P3HP content (heavy grey) were presented. The experiment was carried out in shaking flask in triplicate. Ec, Escherichia coli; Bs, Bacillus subtilis; Pf, Pseudomonas fluorescens; Cg, Corynebacteria glutamicum.
Fig 4.
Effect of the host strain on P3HP production.
Different host strains carrying plasmids pHP302 and pFS03 were grown in minimal medium, and the P3HP production (white), CDW (light grey), and P3HP content (heavy grey) were presented. The experiments were perfomed in triplicate in shake-flask cultures.
Table 2.
Effect of IPTG on the biomass and P3HP production.
Fig 5.
Effect of organic nitrogen on P3HP production.
The strain E. coli JM109(DE3) harboring pHP302 and pFS03 was used, and 3 g/L of soya peptone, yeast extract, beef extract, and tryptone was added into the culture, respectively. Incubation without organic nitrogen was used as the control (CK). The P3HP production (white), CDW (light grey), and P3HP content (heavy grey) were presented. All shake-flask experiments were incubated at 30°C for 48 h after the first time of induction. The experiments were performed in triplicate shake-flask cultures.
Fig 6.
Time profiles for CDW, P3HP production and substrate consumption during an aerobic fed-batch fermentation of E. coli JM109(DE3)/pHP302/pFS03.
The content of P3HP was calculated using the ratio of P3HP weight to cell dry weight.