Bioprocess optimization of interferon β-1-a in Pichia pastoris and its improved inhibitory effect against hepatocellular carcinoma cells
DOI:
https://doi.org/10.1590/s2175-97902022e18984Keywords:
Interferon-β-1a, Pichia pastoris, Expression, Cell survival, Hepatocellular carcinomaAbstract
Interferon-β-1a (INF-β-1a) has gained significant attention due to its emerging applications in the treatment of different human diseases. Therefore, many researchers have attempted to produce it in large quantities and also in a biologically active form using different expression systems. In the present study, we aimed to improve the expression level of INF-β-1a by Pichia pastoris using optimization of culture conditions. The codon-optimized INF-β- 1a gene was cloned into pPICZαA plasmid under the control of alcohol oxidase I (AOX1) promoter. The protein expression was induced using different concentrations of methanol at different pHs and temperatures. The biological activity of produced protein was evaluated by anti-proliferative assay. The ideal culture conditions for the expression of INF-β-1a by P. pastoris were found to be induction with 2% methanol at pH 7.0 culture medium at 30 C which yielded a concentration of 15.5 mg/L INF-β-1a in a shake flask. Our results indicate that differences in glycosylation pattern could result in different biological activities as INF- β-1a produced by P. pastoris could significantly more reduce the cell viability of HepG-2 cells, a hepatocellular carcinoma cell line, than a commercially available form of this protein produced by CHO.
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Ahmad M, Hirz M, Pichler H, Schwab H. Protein expression in Pichia pastoris: recent achievements and perspectives for heterologous protein production. Appl Microbiol Biotechnol. 2014;98(12):5301-5317.
Akbari V, Sadeghi HMM, Jafarian-Dehkordi A, Abedi D, Chou CP. Improved biological activity of a single chain antibody fragment against human epidermal growth factor receptor 2 (HER2) expressed in the periplasm of Escherichia coli. Protein Expression Purif. 2015;116:66-74.
Akbari V, Sadeghi HMM, Jafrian-Dehkordi A, Abedi D, Chou CP. Functional expression of a single-chain antibody fragment against human epidermal growth factor receptor 2 (HER2) in Escherichia coli. J Ind Microbiol Biotechnol. 2014;41(6):947-956.
Almo SC, Love JD. Better and faster: improvements and optimization for mammalian recombinant protein production. Curr Opin Struct Biol. 2014,26:39-43.
Asada H, Uemura T, Yurugi-Kobayashi T, Shiroishi M, Shimamura T, Tsujimoto H, et al. Evaluation of the Pichia pastoris expression system for the production of GPCRs for structural analysis. Microb Cell Fact. 2011;10(1):24.
Baneyx F. Recombinant protein expression in Escherichia coli. Curr Opin biotechnol. 1999;10(5):411-421.
Boettner M, Prinz B, Holz C, Stahl U, Lang C. High-throughput screening for expression of heterologous proteins in the yeast Pichia pastoris. J Biotechnol. 2002;99(1):51-62.
Damasceno LM, Huang C-J, Batt CA. Protein secretion in Pichia pastoris and advances in protein production. Appl Microbiol Biotechnol . 2012;93(1):31-39.
Damdinsuren B, Nagano H, Wada H, Kondo M, Ota H, Nakamura M, et al. Stronger growth-inhibitory effect of interferon (IFN)-beta compared to IFN-alpha is mediated by IFN signaling pathway in hepatocellular carcinoma cells. Int J Oncol. 2007;30(1):201-208.
Dean N. Asparagine-linked glycosylation in the yeast Golgi. Biochim Biophys Acta. 1999;1426(2):309-322.
Demolder J, Fiers W, Contreras R. Human interferon-beta, expressed in Saccharomyces cerevisiae, is predominantly directed to the vacuoles. Influence of modified co- expression of secretion factors and chaperones. J Biotechnol. 1994;32(2):179-189.
Eissazadeh S, Moeini H, Dezfouli MG, Heidary S, Nelofer R, Abdullah MP. Production of recombinant human epidermal growth factor in Pichia pastoris. Braz J Microbiol. 201748(2):286-293.
Han C, Su L, Hong R, Wu S, Wu J. A comparative study of maltooligosyltrehalose synthase from Sulfolobus acidocaldarius expressed in Pichia pastoris and Escherichia coli. Process Biochem. 2017;60:35-41.
Hotter DF, Kirchhoff F. Interferons and beyond: Induction of antiretroviral restriction factors. J Leukocyte Biol. 2018;103(3):465-477.
Hu H, Gao J, He J, Yu B, Zheng P, Huang Z, Mao X, Yu J, Han G, Chen D. Codon optimization significantly improves the expression level of a keratinase gene in Pichia pastoris. PLoS One. 2013;8(3):e58393.
Jakimovski D, Kolb C, Ramanathan M, Zivadinov R, Weinstock-Guttman B. Interferon β for Multiple Sclerosis. Cold Spring Harbor Perspect Med. 2018;a032003.
Kaskow BJ, Baecher-Allan C. Effector T Cells in multiple sclerosis. Cold Spring Harbor Perspect Med. 2018;a029025.
Kruger NJ. The Bradford method for protein quantitation. The protein protocols handbook, Springer: 2002;15-21.
Lassmann H. Multiple sclerosis pathology. Cold Spring Harbor Perspect Med. 2018;8(3):a028936.
Lilie H, Schwarz E, Rudolph R. Advances in refolding of proteins produced in E. coli. Curr Opin Biotechnol. 1998;9(5):497-501.
Macauley-Patrick S, Fazenda ML, McNeil B, Harvey LM. Heterologous protein production using the Pichia pastoris expression system. Yeast. 2005;22(4):249-270.
Medrano RF, Hunger A, Mendonça SA, Barbuto JAM, Strauss BE. Immunomodulatory and antitumor effects of type I interferons and their application in cancer therapy. Oncotarget. 2017;8(41):71249.
Mobasher MA, Ghasemi Y, Montazeri-Najafabady N, Tahzibi A. Expression of recombinant IFN beta 1-b: a comparison between soluble and insoluble forms. Minerva Biotecnologica. 2016;28(1):39-43.
Morowvat MH, Babaeipour V, Rajabi-Memari H, Vahidi H, Maghsoudi N. Overexpression of recombinant human beta interferon (rhINF-β) in periplasmic space of Escherichia coli. Iran J Pharm Res. 2014;13(Suppl):151.
Ogasawara S, Yano H, Momosaki S, Akiba J, Nishida N, Kojiro S, et al. Growth inhibitory effects of IFN-beta on human liver cancer cells in vitro and in vivo. J Interferon Cytokine Res. 2007;27(6):507-516.
Rodriguez J, Spearman M, Huzel N, Butler M. Enhanced production of monomeric interferon-beta by CHO cells through the control of culture conditions. Biotechnol Prog. 2005;21(1):22-30.
Runkel L, Meier W, Pepinsky RB, Karpusas M, Whitty A, Kimball K, et al. Structural and functional differences between glycosylated and non-glycosylated forms of human interferon-β (IFN-β). Pharm Res. 1998;15(4):641-649.
Schmidt F. Recombinant expression systems in the pharmaceutical industry. Applied Microbiol Biotechnol. 2004;65(4):363-372.
Skoko N, Argamante B, Grujičić NK, Tisminetzky SG, Glišin V, Ljubijankić G. Expression and characterization of human interferon-β1 in the methylotrophic yeast Pichia pastoris. Biotechnol Appl Biochem. 2003;38(3):257-265.
Song K, Yoon I-S, Kim NA, Kim D-H, Lee J, Lee HJ, et al. Glycoengineering of interferon-β 1a improves its biophysical and pharmacokinetic properties. PLoS ONE. 2014;9(5):e96967.
Utsumi J, Matsuo-Ogawa E, Nagahata T, Kasama K, Kagawa Y, et al. Carbohydrate-dependent biological activities of glycosylated human interferon-beta on human hepatoblastoma cells in vitro. Microbiol Immunol. 1995;39(1):81-86.
Von Kalckreuth V, Lohse AW, Schramm C. Unmasking autoimmune hepatitis under immunomodulatory treatment of multiple sclerosis-not only beta interferon. Am J Gastroenterol. 2008;103(8):2147.
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