title: De novo production of the flavonoid naringenin in engineered Saccharomyces cerevisiae.
abstract: BACKGROUND
  Flavonoids comprise a large family of secondary plant metabolic intermediates that exhibit a wide variety of antioxidant and human health-related properties. Plant production of flavonoids is limited by the low productivity and the complexity of the recovered flavonoids. Thus to overcome these limitations, metabolic engineering of specific pathway in microbial systems have been envisaged to produce high quantity of a single molecules.
  
  
  RESULTS
  Saccharomyces cerevisiae was engineered to produce the key intermediate flavonoid, naringenin, solely from glucose. For this, specific naringenin biosynthesis genes from Arabidopsis thaliana were selected by comparative expression profiling and introduced in S. cerevisiae. The sole expression of these A. thaliana genes yielded low extracellular naringenin concentrations (<5.5 μM). To optimize naringenin titers, a yeast chassis strain was developed. Synthesis of aromatic amino acids was deregulated by alleviating feedback inhibition of 3-deoxy-d-arabinose-heptulosonate-7-phosphate synthase (Aro3, Aro4) and byproduct formation was reduced by eliminating phenylpyruvate decarboxylase (Aro10, Pdc5, Pdc6). Together with an increased copy number of the chalcone synthase gene and expression of a heterologous tyrosine ammonia lyase, these modifications resulted in a 40-fold increase of extracellular naringenin titers (to approximately 200 μM) in glucose-grown shake-flask cultures. In aerated, pH controlled batch reactors, extracellular naringenin concentrations of over 400 μM were reached.
  
  
  CONCLUSIONS
  The results reported in this study demonstrate that S. cerevisiae is capable of de novo production of naringenin by coexpressing the naringenin production genes from A. thaliana and optimization of the flux towards the naringenin pathway. The engineered yeast naringenin production host provides a metabolic chassis for production of a wide range of flavonoids and exploration of their biological functions.
authors.affiliation: Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC, Delft, The Netherlands.
authors.forename: Frank
authors.initials: F
authors.lastname: Koopman
authors.forename: Jules
authors.initials: J
authors.lastname: Beekwilder
authors.forename: Barbara
authors.initials: B
authors.lastname: Crimi
authors.forename: Adele
authors.initials: A
authors.lastname: van Houwelingen
authors.forename: Robert D
authors.initials: RD
authors.lastname: Hall
authors.forename: Dirk
authors.initials: D
authors.lastname: Bosch
authors.forename: Antonius J A
authors.initials: AJ
authors.lastname: van Maris
authors.forename: Jack T
authors.initials: JT
authors.lastname: Pronk
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Daran
mesh_terms: D010723:3-Deoxy-7-Phosphoheptulonate Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism; D000217:Acyltransferases / Q000235:genetics / Q000378:metabolism; D000642:Ammonia-Lyases / Q000235:genetics / Q000378:metabolism; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics; D002262:Carboxy-Lyases / Q000037:antagonists & inhibitors / Q000378:metabolism; D044950:Flavanones / Q000096:biosynthesis*; D005419:Flavonoids / Q000096:biosynthesis; D060847:Metabolic Engineering; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
chemical_list: D044950:Flavanones; D005419:Flavonoids; D010940:Plant Proteins; D029701:Saccharomyces cerevisiae Proteins; D000217:Acyltransferases; C020141:flavanone synthetase; D010723:3-Deoxy-7-Phosphoheptulonate Synthase; D002262:Carboxy-Lyases; C109058:phenylpyruvate decarboxylase; D000642:Ammonia-Lyases; C023208:L-tyrosine ammonia-lyase; C005273:naringenin