The influence of carbon sources on recombinant-human-growth-hormone production by Pichia pastoris is dependent on phenotype: a comparison of Mut(s) and Mut(+) strains

Orman, Mehmet Ali
Çalık, Pınar
Ozdamar, Tuncer H.
The influence of carbon sources on rhGH (recombinant human growth hormone) production by two Pichia pastoris strains having different methanol utilization phenotypes (P. pastoris-hGH-Mut(+) and P. pastoris-hGH-Mut(s)) was investigated using batch bioreactors. The effect of methanol concentration (C-MeOH) in defined and complex media, and further glycerol/methanol mixed defined media, was analysed systematically over a wide range. With methanol as the sole carbon source, strain Mut(s) grew only slightly, whereas with Mut(+), a cell concentration (C-x) of 6.0 g of dry cells/dm(3) was obtained and an rhGH concentration (C-rhGH) of 0.032 g/dm(3) was produced. In complex medium without glycerol at a C-MeOH of 2% (v/v), a C-rhGH of 0.16 g of rhGH/dm(3) was produced by Mut(s), a value 3-fold higher than that produced by Mut(+), despite the fact that the C-x of Mut(+) (6.1 g/dm(3)) was 2-fold higher than that of Mut(s) (3.0 g/dm(3)). In a glycerol/methanol mixed defined medium, methanol consumption began when glycerol was totally depleted, indicating that glycerol is a repressor of the AOXI (alcohol oxidase-1 gene) promoter. With strain Mut(s) at a glycerol concentration (C-Gly) of 30 g/dm(3) and a C-MeOH of 1% (v/v), the C-rhGH produced was 0.11 g/dm(3), whereas, with the Mut(+) strain, a C-rhGH of 0.06 g/dm(3) was obtained at a C-Gly of 30 g/dm(3) and a C-MeOH of 4%. As methanol is not consumed by Mut(s) strain effectively and the presence of methanol in the fermentation broth triggers induction of the AOXI promoter, our results encourage the use of the Mut(s) strain for rhGH production. In addition to rhGH production, the specific cell growth rates, specific methanol and/or glycerol utilization rates and maintenance coefficients in methanol- and glycerol-based defined media were determined. With a methanol-based defined medium and using the MW strain, a higher specific growth rate (mu) of approx. 0.14 h(-1) was observed during the exponential cell growth phase at a C-MeOH of <= 2.0%. When glycerol was used as a sole carbon source, both phenotypes showed similar cell-growth and glycerol utilization rates. The results of the present study should enable one to optimize the expression of other therapeutic proteins by P. pastoris.


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Influences of the concentration and addition time of alanine-group amino acids, i.e. alanine, leucine and valine, on serine alkaline protease (SAP) synthesis were investigated by Bacillus licheniformis (DSM 1969) carrying pHV1431::subC in a defined medium to identify the amino acids creating intracellular reaction-rate limitation in SAP production. While the precursors of alanine-group amino acids, pyruvate and alanine, did not affect SAP production considerably within the range 0-15 mM, the addition of leu...
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This study aimed to investigate the release of cefuroxime axetil (CF) and calcium from poly(epsilon-caprolactone) (PCL)-calcium sulfate (CaS) implants (PCL:CaS 2:1-10% CF; PCL:CaS 2:1-20% CF; PCL:CaS 1:1-10% CF) for treating infectious bone diseases. Bioactivity, crystallinity and strength, and release profiles under standard and pressurized release conditions were studied. PCL:CaS 2:1-20% CF had slower release than 10% loading. These groups had no significant change in CF and Ca release in response to pres...
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BASALP, A; Özcengiz, Gülay; ALAEDDINOGLU, NG (Springer Science and Business Media LLC, 1992-03-01)
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Rate limiting reactions in the aromatic-group amino acid pathway (AAAP) in Bacillus subtilis for l-phenylalanine (Phe) production were determined as reported elsewhere (Özçelik-Şenver et al., 2004). The biochemical reactions in the AAAP start with the reaction using phosphoenolpyruvate (PEP) and erythrose 4-phosphate (E4P) synthesised in the glycolysis pathway and the PPP, respectively, leading to the formation of Phe via chorismate by branching at prephenate. The branch-point metabolites E4P supplied in vi...
Citation Formats
M. A. Orman, P. Çalık, and T. H. Ozdamar, “The influence of carbon sources on recombinant-human-growth-hormone production by Pichia pastoris is dependent on phenotype: a comparison of Mut(s) and Mut(+) strains,” BIOTECHNOLOGY AND APPLIED BIOCHEMISTRY, pp. 245–255, 2009, Accessed: 00, 2020. [Online]. Available: