In this study, a combined strategy was used to improve the production of Thermomyces dupontii lipase (TDL) in Pichia pastoris. First, the native gene of TDL was optimized based on the codon usage of P. pastoris, ligated to pPICZαA and transformed in P. pastoris X33. A recombinant strain designated X33-T23 with the highest activity (1020 U/mL in shake flasks) amongst 216 recombinant colonies was selected for further investigations. To further increase the production of TDL, nine different secretion helper factor genes were transformed in the recombinant strain, X33-T23. The recombinant strain co-expression with the gene encoding protein disulfide isomerase, designated X33-T23-PDI, exhibited the highest activity in shake flasks (1760 U/mL) and in 5 L bioreactor (57521 U/mL) which were 1.67- and 1.46-fold higher, respectively, than for strain X33-T23. Additionally, the optimization of the inducers (temperature and pH) for the recombinant strain X33-T23-PDI in 5 L bioreactor produced, as expected, much higher lipase activity (81203 U/mL). The results of this study will provide an effective method to produce TDL and give some clues on how to improve production of heterologous proteins in P. pastoris.
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This work was supported by the National High Technology Project of P.R. China (No. 2014AA093514).
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The authors declare that there is no conflict of interest.
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Wang, J., Wu, Z., Zhang, T. et al. High-level expression of Thermomyces dupontii thermophilic lipase in Pichia pastoris via combined strategies. 3 Biotech 9, 62 (2019). https://doi.org/10.1007/s13205-019-1597-8
- Codon optimization
- Pichia pastoris
- Secretion helper factor
- Thermophilic lipase
- Thermomyces dupontii