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Oxygen-limited control of methanol uptake for improved production of a single-chain antibody fragment with recombinant Pichia pastoris

  • Biotechnological Products and Process Engineering
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Abstract

The yeast Pichia pastoris is a suitable production system for recombinant proteins due to its strong methanol-inducible AOX1 promoter. A key parameter of the production process is the specific methanol uptake rate. To control the methanol uptake and simultaneously maintain a constant methanol concentration during the production phase, two strategies were developed to generate purposeful oxygen limitation and to feed-forward control the specific methanol uptake rate into the optimum range. First, the cell density at induction was adjusted by prolonged preinduction glycerol feeding. Alternatively, the airflow rate was restricted and increased in parallel with the biomass. While the product accumulation started 20 h earlier with the first approach, the specific production rate of a single-chain antibody fragment was three times higher in the latter case. After 70 h of production, both schemes yielded product concentrations in the gram-per-liter range. Moreover, they release the requirement for dosage of pure oxygen and thereby can facilitate the scale-up of the production process. The different production profiles indicate that the impact of specific methanol uptake rate on protein production by recombinant P. pastoris depends on the control mode.

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Authors and Affiliations

  1. Department of Biochemistry/Biotechnology, Institute for Biotechnology, Martin-Luther-University Halle-Wittenberg, Kurt-Mothes-Straße 3, 06120, Halle, Germany

    Narendar K. Khatri & Frank Hoffmann

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  1. Narendar K. Khatri
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  2. Frank Hoffmann
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Correspondence to Frank Hoffmann.

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Khatri, N.K., Hoffmann, F. Oxygen-limited control of methanol uptake for improved production of a single-chain antibody fragment with recombinant Pichia pastoris . Appl Microbiol Biotechnol 72, 492–498 (2006). https://doi.org/10.1007/s00253-005-0306-1

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  • DOI: https://doi.org/10.1007/s00253-005-0306-1

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