Fig 1.
Metabolic pathway involved in the conversion of propionate to 3-hydroxypropionate in recombinant E. coli strains.
The enzymes in bold are overexpressed, while disrupted pathway steps are indicated by the bold “×” symbols. MC cycle: methylcitrate cycle; TCA cycle: tricarboxylic acid cycle; PCT: propionate CoA-transferase; PACD: propionyl-CoA dehydrogenase; HPCD: 3-hydroxypropionyl-CoA dehydratase; YgfH: propionyl-CoA: succinate-CoA transferase; PrpC: methylcitrate synthase.
Fig 2.
Construction of plasmids pET-PH and pACYCDuet-PACD.
Table 1.
Bacterial strains and plasmids used in the study.
Fig 3.
SDS-PAGE analysis of cell extract of the host E. coli, Ec-PPH and Ec-P.
(Lane 1) protein standard, (Lane 2) host E. coli, (Lane 3) Ec-PPH expressing PACD, PCT and HPCD, (Lane 4) Ec-P with PACD expression alone. The arrow marks indicate the corresponding target proteins.
Fig 4.
Time-course profile of propionic acid consumption and production of 3-HP for two strains, Ec-PPH and Ec-P.
(A) Ec-PPH (expressing PACD PCT and HPCD, closed symbols) and Ec-P (expressing PACD, open symbols) were cultured in MI medium at 30°C. (B) Ec-PPH was cultivated in medium with glucose (1%, closed symbols) and medium without glucose (open symbols) at 30°C. (C) Ec-PPH cultivation in MI medium at 30°C (closed symbols) and 37°C (open symbols). Symbols: 3-HP (circle), propionic acid (triangle). The experiment was done in triplicate. The error bars show standard errors and are not shown if the error did not exceed the size of the symbol.
Fig 5.
Production of 3-HP by recombinant E. coli deletion mutants expressing pacd, pct and hpcd.
(A) WT Ec, (B) Ec-ΔygfH, (C) Ec-ΔprpC, and (D) Ec-ΔygfH ΔprpC. Symbols: 3-HP (square), cell mass biomass (circle), pH (triangle). The error bars denote standard errors of the mean from triplicate flasks and are not shown where the error did not exceed the size of the symbol.