Metabolic engineering of Pseudomonas mendocina NK-01 for enhanced production of medium-chain-length polyhydroxyalkanoates with enriched content of the dominant monomer

Abstract

In this study, six genes involved in β-oxidation pathway of P. mendocina NK-01 were deleted to construct mutant strains NKU-∆β1 and NKU-∆β5. Compared with the wild strain NKU, the mcl-PHA titers of NKU-∆β5 were respectively increased by 5.58- and 4.85-fold for culturing with sodium octanoate and sodium decanoate. And the mcl-PHA titers of NKU-∆β1 was increased by 10.02-fold for culturing with dodecanoic acid. The contents of dominant monomers 3-hydroxydecanoate (3HD) and 3-hydroxydodecanoate (3HDD) of the mcl-PHA synthesized by NKU-∆β5 were obviously increased to 90.01 and 58.60 mol%, respectively. Further deletion of genes phaG and phaZ, the 3HD and 3HDD contents were further improved to 94.71 and 68.67 mol%, respectively. The highest molecular weight of mcl-PHA obtained in this study was 80.79 × 10⁴ Da, which was higher than the previously reported mcl-PHA. With the increase of dominant monomer contents, the synthesized mcl-PHA showed better thermal properties, mechanical properties and crystallization properties. Interestingly, the cell size of NKU-∆β5 was larger than that of NKU due to the accumulation of more PHA granules. This study indicated that a systematically metabolic engineering approach for P. mendocina NK-01 could significantly improve the mcl-PHA titer, dominant monomer contents and physical properties of mcl-PHA.

Keywords: Metabolic engineering; Pseudomonas mendocina; mcl-PHA.