Objectives: Different cultivation conditions and parameters were evaluated to improve the production and secretion of a recombinant Phanerochaete chrysosporium lipH8 gene in Komagataella phaffii (Pichia pastoris).
Results: The recombinant lipH8 gene with its native secretion signal was successfully cloned and expressed in Komagataella phaffii (Pichia pastoris) under the control of the alcohol oxidase 1 promoter (PAOX1). The results revealed that co-feeding with sorbitol and methanol increased rLiP secretion by 5.9-fold compared to the control conditions. The addition of 1 mM FeSO4 increased LiP activity a further 6.0-fold during the induction phase. Moreover, the combination of several optimal conditions and parameters yielded an extracellular rLiP activity of 20.05 U l-1, which is more than ten-fold higher relative to standard growth conditions (BMM10 medium, pH 6 and 30 °C).
Conclusion: Extracellular activity of a recombinant LiP expressed in P. pastoris increased more than ten-fold when co-feeding sorbitol and methanol as carbon sources, together with urea as nitrogen source, FeSO4 supplementation, lower pH and lower cultivation temperature.
Keywords: Co-carbon feeding; Lignin peroxidase; Medium optimization; Pichia pastoris; Recombinant enzymes.
© 2022. The Author(s), under exclusive licence to Springer Nature B.V.