Biosynthesis of isocitric acid by the yeast Yarrowia lipolytica and its regulation

We studied the biosynthesis of isocitric acid from rapeseed (canola) oil by the yeast Yarrowia lipolytica and its regulation. We determined a fundamental possibility for directed biosynthesis of isocitric acid by Y. lipolytica yeast, with only minimal amounts of citric acid byproduct, when grown on a medium containing canola oil. Wild type strains of Y. lipolytica were mutagenized by UV irradiation and treatment with N-methyl-N'-nitro-N-nitrosoguanidine (NG). Subsequent selection on media with acetate and isocitrate resulted in isolation of a mutant Y. lipolytica UV/NG that synthesized isocitrate and citrate at a ratio of 2.7 : 1. In the parent strain, this ratio is 1 : 1. Inhibition of isocitrate lyase, a key enzyme in the metabolism of isocitric acid, by the addition of itaconic acid resulted in increased synthesis of isocitrate, with a ratio of isocitrate to citrate reaching 6 : 1. The cultivation of the Y. lipolytica UV/NG mutant in a pilot industrial fermenter in the presence of itaconic acid resulted in the production of 88.7 g/L of isocitric acid with a yield of 90%.