Yarrowia lipolyticachassis strains engineered to produce aromatic amino acids via the shikimate pathway

Abstract

Yarrowia lipolytica is widely used as a microbial producer of lipids and lipid derivatives. Here, we exploited this yeast's potential to generate aromatic amino acids by developing chassis strains optimized for the production of phenylalanine, tyrosine and tryptophan. We engineered the shikimate pathway to overexpress a combination of Y. lipolytica and heterologous feedback-insensitive enzyme variants. Our best chassis strain displayed high levels of de novo Ehrlich metabolite production (up to 0.14 g l À1 in minimal growth medium), which represented a 93fold increase compared to the wild-type strain (0.0015 g l À1 ). Production was further boosted to 0.48 g l À1 when glycerol, a low-cost carbon source, was used, concomitantly to high secretion of phenylalanine precursor (1 g l À1 ). Among these metabolites, 2-phenylethanol is of particular interest due to its rose-like flavour. We also established a production pathway for generating protodeoxyviolaceinic acid, a dye derived from tryptophan, in a chassis strain optimized for chorismate, the precursor of tryptophan. We have thus demonstrated that Y. lipolytica can serve as a platform for the sustainable de novo bioproduction of high-value aromatic compounds, and we have greatly improved our understanding of the potential feedback-based regulation of the shikimate pathway in this yeast.

Bioproduction of 2-Phenylethanol through Yeast Fermentation on Synthetic Media and on Agro-Industrial Waste and By-Products: A Review

“…Particularly, dedicated genetic toolboxes can be expanded in order to artificially generate non-native metabolites [ 78 ]. For instance, different strategies have been developed in order to allow engineered Y. lipolytica to grow on various cheap substrates, to generate aromatic compounds including 2-PE, and to facilitate product extraction [ 79 ].…”

Engineering ergothioneine production in Yarrowia lipolytica

“…Lipid production requires cytosolic acetyl‐CoA, malonyl‐CoA, and NADPH [ 11 ]. As these precursors are also required for isoprenoids, polyketides, and other metabolites, Y. lipolytica is also a promising cell factory for the production of such metabolites [ 11 , 12 , 13 , 14 , 15 ]. Yarrowia lipolytica has been engineered for the production of β‐carotene [ 14 ], β‐farnesene, limonene, valencene, squalene, 2,3‐oxidosqualene [ 12 ], aromatic amino acids [ 15 ], and resveratrol [ 13 ].…”

“…As these precursors are also required for isoprenoids, polyketides, and other metabolites, Y. lipolytica is also a promising cell factory for the production of such metabolites [ 11 , 12 , 13 , 14 , 15 ]. Yarrowia lipolytica has been engineered for the production of β‐carotene [ 14 ], β‐farnesene, limonene, valencene, squalene, 2,3‐oxidosqualene [ 12 ], aromatic amino acids [ 15 ], and resveratrol [ 13 ]. Additionally, Y. lipolytica is used for the production of lipases, β‐mannases, laccases, amylases, and proteases [ 16 ].…”

Metabolic profiles and fingerprints for the investigation of the influence of nitisinone on the metabolism of the yeast Saccharomyces cerevisiae

“…One path yields tryptophan while the other, via a Claisen rearrangement, yields prephenate, with is ultimately transformed into phenylalanine and tyrosine 22 . The tryptophan biosynthesis involves ve steps: it starts with the conversion of chorismate to anthranilate, which is converted to TRF in four sequential steps by enzymes 23 . Tyrosine can be formed from arogenate or 4-hydroxyphenyl pyruvate, whereas phenylalanine may originate from prephenate passing through either arogenate or phenylpyruvate as intermediates 22 .…”