| Summary: | Poly(3-hydroxybutyrate-<i>co</i>-3-hydroxypropionate) [P(3HB-<i>co</i>-3HP)] is a biodegradable and biocompatible polyester with improved and expanded material properties compared with poly(3-hydroxybutyrate) (PHB). This study engineered a robust malonyl-CoA pathway in <i>Cupriavidus necator</i> for the efficient supply of a 3HP monomer, and could achieve the production of [P(3HB-<i>co</i>-3HP)] from variable oil substrates. Flask level experiments followed by product purification and characterization found the optimal fermentation condition (soybean oil as carbon source, 0.5 g/L arabinose as induction level) in general consideration of the PHA content, PHA titer and 3HP molar fraction. A 5 L fed-batch fermentation (72 h) further increased the dry cell weight (DCW) to 6.08 g/L, the titer of [P(3HB-<i>co</i>-3HP)] to 3.11 g/L and the 3HP molar fraction to 32.25%. Further improving the 3HP molar fraction by increasing arabinose induction failed as the engineered malonyl-CoA pathway was not properly expressed under the high-level induction condition. With several promising advantages (broader range of economic oil substrates, no need for expensive supplementations such as alanine and VB<sub>12</sub>), this study indicated a candidate route for the industrial level production of [P(3HB-<i>co</i>-3HP)]. For future prospects, further studies are needed to further improve the strain and the fermentation process and expand the range of relative products.
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