Design of an efficient medium for heterologous protein production in Yarrowia lipolytica: case of human interferon alpha 2b

Microbial Cell Factories

Research

Jean-Marc Nicaud¹  Atef Ayed²  Héla Kallel²  Najla Gasmi³ 

[1]INRA, UMR1319 Micalis, Domaine de Vilvert, Jouy-en-Josas, F-78352, France

[2]CNRS, UMR1319 Micalis, Domaine de Vilvert, Jouy-en-Josas, F-78352, France

[3]Unité de Biofermentation, Institut Pasteur Tunis, 13, place Pasteur. BP 74, 1002, Tunis, Tunisie

[4]Unité de Biofermentation, Institut Pasteur Tunis, 13, place Pasteur. BP 74, 1002, Tunis, Tunisie

[5]INRA, UMR1319 Micalis, Domaine de Vilvert, Jouy-en-Josas, F-78352, France

关键词: Oleic Acid;    Heterologous Protein;    Pichia Pastoris;    Lipid Body;    Recombinant Protein Production;   

DOI  :  10.1186/1475-2859-10-38

 received in 2010-12-14, accepted in 2011-05-20,  发布年份 2011

来源: Springer

【 摘 要 】

BackgroundThe non conventional yeast Yarrowia lipolytica has aroused a strong industrial interest for heterologous protein production. However most of the studies describing recombinant protein production by this yeast rely on the use of complex media, such media are not convenient for large scale production particularly for products intended for pharmaceutical applications. In addition medium composition can also affect the production yield. Hence it is necessary to design an efficient medium for therapeutic protein expression by this host.ResultsFive different media, including four minimal media and a complex medium, were assessed in shake flasks for the production of human interferon alpha 2b (hIFN α2b) by Y. lipolytica under the control of POX2 promoter inducible with oleic acid. The chemically defined medium SM4 formulated by Invitrogen for Pichia pastoris growth was the most suitable. Using statistical experimental design this medium was further optimized. The selected minimal medium consisting in SM4 supplemented with 10 mg/l FeCl3, 1 g/l glutamate, 5 ml/l PTM1 (Pichia Trace Metals) solution and a vitamin solution composed of myo-inositol, thiamin and biotin was called GNY medium. Compared to shake flask, bioreactor culture in GNY medium resulted in 416-fold increase of hIFN α2b production and 2-fold increase of the biological activity.Furthermore, SM4 enrichment with 5 ml/l PTM1 solution contributed to protect hIFN α2b against the degradation by the 28 kDa protease identified by zymography gel in culture supernatant. The screening of the inhibitory effect of the trace elements present in PTM1 solution on the activity of this protease was achieved using a Box-Behnken design. Statistical data analysis showed that FeCl3 and MnSO4 had the most inhibitory effect.ConclusionWe have designed an efficient medium for large scale production of heterologous proteins by Y. lipolytica. The optimized medium GNY is suitable for the production of hIFN α2b with the advantage that no complex nitrogen sources with non-defined composition were required.

【 授权许可】

Unknown   
© Gasmi et al; licensee BioMed Central Ltd. 2011. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.