Roles of multiple acyl-CoA oxidases in the routing of carbon flow towards β-oxidation and polyhydroxyalkanoate biosynthesis in Yarrowia lipolytica

Abstract

The oleaginous yeast Yarrowia lipolytica possesses six acyl-CoA oxidase (Aox) isoenzymes encoded by genes POX1-POX6. The respective roles of these multiple Aox isoenzymes were studied in recombinant Y. lipolytica strains that express heterologous polyhydroxyalkanoate (PHA) synthase (phaC) of Pseudomonas aeruginosa in varying POX genetic backgrounds, thus allowing assessment of the impact of specific Aox enzymes on the routing of carbon flow to β-oxidation or to PHA biosynthesis. Analysis of PHA production yields during growth on fatty acids with different chain lengths has revealed that the POX genotype significantly affects the PHA levels, but not the monomer composition of PHA. Aox3p function was found to be responsible for 90% and 75% of the total PHA produced from either C9:0 or C13:0 fatty acid, respectively, whereas Aox5p encodes the main Aox involved in the biosynthesis of 70% of PHA from C9:0 fatty acid. Other Aoxs, such as Aox1p, Aox2p, Aox4p and Aox6p, were not found to play a significant role in PHA biosynthesis, independent of the chain length of the fatty acid used. Finally, three known models of β-oxidation are discussed and it is shown that a 'leaky-hose pipe model' of the cycle can be applied to Y. lipolytica.

Exploring medium-chain-length polyhydroxyalkanoates production in the engineered yeast Yarrowia lipolytica

“…Thereafter, mcl-PHA was similarly synthesized in peroxisomes of Pichia pastoris, and reached 1 % DCW using oleic acid as substrate [24]. Recently, Y. lipolytica was engineered for synthesis of mcl-PHA, which was further improved by inactivating the R-3-hydroxyacyl-CoA dehydrogenase domain of MFE2 and blocking the neutral lipid synthesis pathway using 0.2 % tridecanoic acid in YNB minimal medium [10,11]. These studies indicated that Y. lipolytica, which exhibits efficient lipid/fatty acid catabolism and anabolism capacities, owns a tremendous potential for mcl-PHA synthesis in comparison with other yeasts.…”

“…It has six genes (POX1-POX6) encoding isoenzymes of acyl-CoA oxidases (Aoxs) with different substrate specificities that catalyze the first step and also the limiting step of β-oxidation of fatty acids [5,10]. In contrast, S. cerevisiae has only one Aox-encoding gene.…”

“…Another effectual work was the synthesis of trans-10, cis-12 conjugated linoleic acid (CLA) through heterologously expressing linoleic acid isomerase gene from Propionibacterium acnes, which was further enhanced by a modified promoter and co-expression of a ∆12-desaturase gene [33,34]. Recently, Haddouche et al examined the feasibility of using Y. lipolytica for PHA biosynthesis and investigated the roles of multiple acyl-CoA oxidases in the routing of carbon flow towards β-oxidation [10].…”

Engineering the Saccharomyces cerevisiae β-Oxidation Pathway to Increase Medium Chain Fatty Acid Production as Potential Biofuel

“…In several studies, fatty acids are overproduced in microbes and later converted to fuels or chemicals through chemical conversion [1] or bioconversion [2], [3]. Other studies directly utilize fatty acid-rich feedstock as a carbon source to produce biofuels and chemicals through the complete oxidation of fatty acids [4] or using the intermediates of β-oxidation [5], [6]. The availability of fatty acids from nonedible fatty acid-rich crops, oleaginous fungi/algae and industrial by-products encourage the use of fatty acid feedstock as a cheap carbon source.…”

Yarrowia lipolytica as an Oleaginous Cell Factory Platform for Production of Fatty Acid-Based Biofuel and Bioproducts

“…Although bacteria can produce PHA, the oleaginous yeast platform is also highly competitive, due to its ability to utilize a wider range of cheap feedstock, and intrinsically accumulate lipid and convert it to PHA. For example, the expression of PHA synthetase from Pseudomonas aeruginosa in this yeast under different POX genetic profiles has been reported (Haddouche et al, 2010). Based on the results, only Aox3p and Aox5p have been found to be involved in PHA biosynthesis from specific FAs.…”