Pilot-scale production of peroxygenase from agrocybe aegerita
Fabio Tonin (TU Delft - BT/Biocatalysis)
F. Tieves (TU Delft - BT/Biocatalysis)
Sébastien Jean Paul Willot (TU Delft - BT/Biocatalysis)
Anouska Van Troost (Student TU Delft)
Remco Van Oosten (TU Delft - BT/Biocatalysis)
Stefaan Breestraat (Bioprocess Pilot Facility)
Sander Van Pelt (Bioprocess Pilot Facility)
Miguel Alcalde (Institute of Catalysis, CSIC, Madrid)
Frank Hollmann (TU Delft - BT/Biocatalysis)
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Abstract
The pilot-scale production of the peroxygenase from Agrocybe aegerita (rAaeUPO) is demonstrated. In a fed-batch fermentation of the recombinant Pichia pastoris, the enzyme was secreted into the culture medium to a final concentration of 0.29 g L-1 corresponding to 735 g of the peroxygenase in 2500 L of the fermentation broth after 6 days. Due to nonoptimized downstream processing, only 170 g of the enzyme has been isolated. The preparative usefulness of the so-obtained enzyme preparation has been demonstrated at a semipreparative scale (100 mL) as an example of the stereoselective hydroxylation of ethyl benzene. Using an adjusted H2O2 feed rate, linear product formation was observed for 7 days, producing more than 5 g L-1 (R)-1-phenyl ethanol. The biocatalyst performed more than 340.000 catalytic turnovers (942 g of the product per gram of rAaeUPO).
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