Pichia pastoris versus Saccharomyces cerevisiae: a case study on the recombinant production of human granulocyte-macrophage colony-stimulating factor.

Autor: Tran AM; Faculty of Biology and Biotechnology, University of Science, VNU-HCM, Ho Chi Minh, Vietnam., Nguyen TT; Faculty of Biology and Biotechnology, University of Science, VNU-HCM, Ho Chi Minh, Vietnam., Nguyen CT; Faculty of Biology and Biotechnology, University of Science, VNU-HCM, Ho Chi Minh, Vietnam., Huynh-Thi XM; Faculty of Biology and Biotechnology, University of Science, VNU-HCM, Ho Chi Minh, Vietnam., Nguyen CT; Faculty of Biology and Biotechnology, University of Science, VNU-HCM, Ho Chi Minh, Vietnam., Trinh MT; Faculty of Biology and Biotechnology, University of Science, VNU-HCM, Ho Chi Minh, Vietnam., Tran LT; Faculty of Biology and Biotechnology, University of Science, VNU-HCM, Ho Chi Minh, Vietnam., Cartwright SP; School of Life and Health Sciences, Aston University, Birmingham, UK., Bill RM; School of Life and Health Sciences, Aston University, Birmingham, UK., Tran-Van H; Faculty of Biology and Biotechnology, University of Science, VNU-HCM, Ho Chi Minh, Vietnam. tvhieu@hcmus.edu.vn.
Jazyk: angličtina
Zdroj: BMC research notes [BMC Res Notes] 2017 Apr 04; Vol. 10 (1), pp. 148. Date of Electronic Publication: 2017 Apr 04.
DOI: 10.1186/s13104-017-2471-6
Abstrakt: Background: Recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) is a glycoprotein that has been approved by the FDA for the treatment of neutropenia and leukemia in combination with chemotherapies. Recombinant hGM-CSF is produced industrially using the baker's yeast, Saccharomyces cerevisiae, by large-scale fermentation. The methylotrophic yeast, Pichia pastoris, has emerged as an alternative host cell system due to its shorter and less immunogenic glycosylation pattern together with higher cell density growth and higher secreted protein yield than S. cerevisiae. In this study, we compared the pipeline from gene to recombinant protein in these two yeasts.
Results: Codon optimization in silico for both yeast species showed no difference in frequent codon usage. However, rhGM-CSF expressed from S. cerevisiae BY4742 showed a significant discrepancy in molecular weight from those of P. pastoris X33. Analysis showed purified rhGM-CSF species with molecular weights ranging from 30 to more than 60 kDa. Fed-batch fermentation over 72 h showed that rhGM-CSF was more highly secreted from P. pastoris than S. cerevisiae (285 and 64 mg total secreted protein/L, respectively). Ion exchange chromatography gave higher purity and recovery than hydrophobic interaction chromatography. Purified rhGM-CSF from P. pastoris was 327 times more potent than rhGM-CSF from S. cerevisiae in terms of proliferative stimulating capacity on the hGM-CSF-dependent cell line, TF-1.
Conclusion: Our data support a view that the methylotrophic yeast P. pastoris is an effective recombinant host for heterologous rhGM-CSF production.
Databáze: MEDLINE
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