Role of fatty acid de novo biosynthesis in polyhydroxyalkanoic acid (PHA) and rhamnolipid synthesis by pseudomonads: Establishment of the transacylase (PhaG)-mediated pathway for PHA biosynthesis in Escherichia coli

Since Pseudomonas aeruginosa is capable of biosynthesis of polyhydroxyalkan oic acid (PHA) and rhamnolipids, which contain lipid moieties that are deri ved from fatty acid biosynthesis, we investigated various fab mutants from P. aeruginosa with respect to biosynthesis of PHAs and rhamnolipids, All is ogenic fabA, fabB, fabI rhlG, and phaG mutants from P. aeruginosa showed de creased PHA accumulation and rhamnolipid production. In the phaG (encoding transacylase) mutant rhamnolipid production was only slightly decreased, Ex pression of phaG from Pseudomonas putida and expression of the beta -ketoac yl reductase gene rhlG from P. aeruginosa in these mutants indicated that P haG catalyzes diversion of intermediates of fatty acid de novo biosynthesis towards PHA biosynthesis, whereas RhlG catalyzes diversion towards rhamnol ipid biosynthesis. These data suggested that both biosynthesis pathways are competitive. In order to investigate whether PhaG is the only linking enzy me between fatty acid de novo biosynthesis and PHA biosynthesis, we generat ed five Tn5 mutants of P, pah:da strongly impaired in PW production from gl uconate, AII mutants were complemented by the phaG gene from P. putida, ind icating that the transacylase-mediated PHA. biosynthesis route represents t he only metabolic link between fatty acid de novo biosynthesis and PRA bios ynthesis in this bacterium. The transacylase-mediated PKA biosynthesis rout e from gluconate was established in recombinant E. coli, coexpressing the c lass II PHA. synthase gene phaC1 together with the phaG gene from P. putida , only when fatty acid de novo biosynthesis was partially inhibited by tric losan, The accumulated PHA contributed to 2 to 3% of cellular dry weight.