Enhanced protein production by sorbitol co-feeding with methanol in recombinant Pichia pastoris strains
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Pichia pastoris strains carrying 1, 6, 12, and 18 copies of the porcine insulin precursor (PIP) gene, were employed to investigate the effects of sorbitol co-feeding with methanol on the physiology of the strains. Multicopy clones of the methylotrophic yeast were generated to vary the PIP gene dosage and recombinant proteins. Elevated gene dosage increased levels of the recombinant PIP protein when methanol served as the sole carbon and energy source i.e., an increase of 1.9% for a strain carrying 1 copy, 42.6% for a strain carrying 6 copies, 34.7% for a strain carrying 12 copies and 80.9% for a strain carrying 18 copies, respectively (using sorbitol co-feeding with methanol during the induction phase). However, it had no significant influence on a lower gene dosage strain (1 copy), but this approach affirmed enhancement in cell growth and PIP production for higher gene dosage strain (6, 12, and 18 copies) via using sorbitol co-feeding with methanol. Additionally, the co-feeding strategy could hold vital importance for recombinant protein production by a multi-copy P. pastoris system.
Keywordssorbitol co-feeding multi-copy Pichia pastoris porcine insulin precursor
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- 1.Kim, M. J., S. H. Kim, J. H. Lee, J. H. Seo, J. H. Kim, Y. H. Kim, and S. W. Nam (2008) High-level secretory expression of human procarboxypeptidase B by fed-batch cultivation of Pichia pastoris and its partial characterization. J. Microbiol. Biotechnol. 18: 1938–1944.Google Scholar
- 4.Weidner, M., M. Taupp, and S. J. Hallam (2010) Expression of recombinant proteins in the methylotrophic yeast Pichia pastoris. J. Vis. Exp. 36: 1862.Google Scholar
- 7.Scorer, C. A., J. J. Clare, W. R. McCombie, M. A. Romanos, and K. Sreekrishna (1994) Rapid selection using G418 of high copy number transformants of Pichia pastoris for high-level foreign gene expression. Biotechnology. 12: 181–184.Google Scholar
- 9.Charoenrat, T., K. Sangprapai, P. Promdonkoy, K. Kocharin, S. Tanapongpipat, and N. Roongsawang (2015) Enhancement of thermostable β-glucosidase production in a slow methanol utilization strain of Pichia pastoris by optimization of the specific methanol supply rate. Biotechnol. Bioproc. Eng. 20: 315–323.CrossRefGoogle Scholar
- 13.Mansur, M., C. Cabello, L. Hernandez, J. Pais, L. Varas, J. Valdes, Y. Terrero, A. Hidalgo, L. Plana, V. Besada, L. Garcia, E. Lamazares, L. Castellanos, and E. Martinez (2005) Multiple gene copy number enhances insulin precursor secretion in the yeast Pichia pastoris. Biotechnol. Lett. 27: 339–345.CrossRefGoogle Scholar
- 17.Prabhu, A. A., B. Mandal, and V. V. Dasu (2017) Medium optimization for high yield production of extracellular human interferon-γ from Pichia pastoris: A statistical optimization and neural network-based approach. Kor. J. Chem. Eng. 1–13.Google Scholar
- 22.Athmaram, T. N., S. Saraswat, A. K. Singh, M. K. Rao, N. Gopalan, V. V. Suryanarayana, and P. V. Rao (2012) Influence of copy number on the expression levels of pandemic influenza hemagglutinin recombinant protein in methylotrophic yeast Pichia pastoris. Virus Genes. 45: 440–451.CrossRefGoogle Scholar
- 28.Sreekrishna, K., R. G. Brankamp, K. E. Kropp, D. T. Blankenship, J. T. Tsay, P. L. Smith, J. D. Wierschke, A. Subramaniam, and L. A. Birkenberger (1997) Strategies for optimal synthesis and secretion of heterologous proteins in the methylotrophic yeast Pichia pastoris. Gene. 190: 55–62.CrossRefGoogle Scholar
- 30.Calik, P., B. Bozkurt, G. Zerze, B. Inankur, E. Bayraktar, E. Boy, M. A. Orman, E. Acik, and T. Ozdamar (2013) Effect of cosubstrate sorbitol different feeding strategies on human growth hormone production by recombinant Pichia pastoris. J. Chem. Technol. Biotechnol. 88: 1631–1640.CrossRefGoogle Scholar