Abstract
BackgroundThe oleaginous yeast Yarrowia lipolytica is increasingly used as alternative cell factory for the production of recombinant proteins. At present, several promoters with different strengths have been developed based either on the constitutive pTEF promoter or on oleic acid inducible promoters such as pPOX2 and pLIP2. Although these promoters are highly efficient, there is still a lack of versatile inducible promoters for gene expression in Y. lipolytica.ResultsWe have isolated and characterized the promoter of the EYK1 gene coding for an erythrulose kinase. pEYK1 induction was found to be impaired in media supplemented with glucose and glycerol, while the presence of erythritol and erythrulose strongly increased the promoter induction level. Promoter characterization and mutagenesis allowed the identification of the upstream activating sequence UAS1EYK1. New hybrid promoters containing tandem repeats of either UAS1XPR2 or UAS1EYK1 were developed showing higher expression levels than the native pEYK1 promoter. Furthermore, promoter strength was improved in a strain carrying a deletion in the EYK1 gene, allowing thus the utilization of erythritol and erythrulose as free inducer.ConclusionsNovel tunable and regulated promoters with applications in the field of heterologous protein production, metabolic engineering, and synthetic biology have been developed, thus filling the gap of the absence of versatile inducible promoter in the yeast Y. lipolytica.
Highlights
The oleaginous yeast Yarrowia lipolytica is increasingly used as alternative cell factory for the produc‐ tion of recombinant proteins
We report on the identification of the inducible promoter from the EYK1 gene encoding an erythrulose kinase in Y. lipolytica, the characterisation of its regulatory elements and the development of hybrid derivatives promoters showing different induction strengths and regulatory patterns depending on the genetic background of the recipient strain (WT or ∆eyk1)
EYK1 promoter is induced by erythritol and erythrulose To date, two different pathways have been reported for erythritol catabolism
Summary
The oleaginous yeast Yarrowia lipolytica is increasingly used as alternative cell factory for the produc‐ tion of recombinant proteins. Comparison of strength and regulation of promoters from the glycerol3-phosphate dehydrogenase (G3P), the isocitrate lyase (ICL1) and of genes involved in beta-oxidation pathway such as the 3-oxo-acyl-CoA thiolase (POT1) and the acylCoA oxidases (POX2, POX1 and POX5) was reported [22] This provided the first strong promoters inducible by glycerol (G3P), ethanol (ICL) and oleic acid (POT1 and POX2). Other inducible promoters available in Y. lipolytica are those from genes encoding isocitrate lyase (pICL1, [22]), fructose-bisphosphate aldolase (pFBA1, [27]), phosphoglycerate mutase (pGPM) or glycerol-3-phosphate O-acyltransferase (pGPAT) They have been used for heterologous protein production with various successes (for a review see [1, 28])
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
