title: Development of a cultivation process for the enhancement of human interferon alpha 2b production in the oleaginous yeast, Yarrowia lipolytica.
abstract: BACKGROUND
  As an oleaginous yeast, Yarrowia lipolytica is able to assimilate hydrophobic substrates. This led to the isolation of several promoters of key enzymes of this catabolic pathway. Less is known about the behavior of Y. lipolytica in large bioreactors using these substrates. There is therefore a lack of established know-how concerning high cell density culture protocols of this yeast. Consequently, the establishment of suitable induction conditions is required, to maximize recombinant protein production under the control of these promoters.
  
  
  RESULTS
  Human interferon α2b (huIFN α2b) production in Yarrowia lipolytica was used as a model for the enhancement of recombinant protein production under the control of the oleic acid (OA)-inducible promoter POX2. Cell viability and heterologous protein production were enhanced by exponential glucose feeding, to generate biomass before OA induction. The optimal biomass level before induction was determined (73 g L(-1)), and glucose was added with oleic acid during the induction phase. Several oleic acid feeding strategies were assessed. Continuous feeding with OA at a ratio of 0.02 g OA per g dry cell weight increased huIFNα2b production by a factor of 1.88 (425 mg L(-1)) and decreased the induction time (by a factor of 2.6, 21 h). huIFN α2b degradation by an aspartic protease secreted by Y. lipolytica was prevented by adding pepstatin (10 μM), leading to produce a 19-fold more active huIFN α2b (26.2 × 10(7) IU mg(-1)).
  
  
  CONCLUSIONS
  Y. lipolytica, a generally regarded as safe (GRAS) microorganism is one of the most promising non conventional yeasts for the production of biologically active therapeutic proteins under the control of hydrophobic substrate-inducible promoter.
authors: Unité de Biofermentation, Institut Pasteur Tunis, 13 place Pasteur, BP 74 1002, Tunis, Tunisie.
authors: Najla
authors: N
authors: Gasmi
authors: Atef
authors: A
authors: Ayed
authors: Billel Bel Hadj
authors: BB
authors: Ammar
authors: Rim
authors: R
authors: Zrigui
authors: Jean-Marc
authors: JM
authors: Nicaud
authors: Héla
authors: H
authors: Kallel
mesh_terms: D018533:Biomass; D019149:Bioreactors / Q000382:microbiology; D005818:Genetic Engineering; D005947:Glucose / Q000378:metabolism; D006801:Humans; D007218:Industrial Microbiology / Q000379:methods*; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000096:biosynthesis* / Q000235:genetics; D019301:Oleic Acid / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D025062:Yarrowia / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*
chemical_list: D000077190:Interferon alpha-2; D016898:Interferon-alpha; D011994:Recombinant Proteins; D019301:Oleic Acid; D005947:Glucose