Production Design of Efficient Recombinant Human Interleukin-4 (rhIL-4) under Specific Promoter in Escherichia coli
Trends in Peptide and Protein Sciences,
Vol. 3 (2018),
1 June 2018
,
Page 1-9 (e3)
https://doi.org/10.22037/tpps.v3i0.21996
Abstract
Recombinant DNA technology plays a vital role in improving health conditions by developing new pharmaceuticals. Recently, some of the cytokines as other recombinant proteins could be produced using the recombinant DNA technology. The role of Recombinant IL-4 in allergy, autoimmunity, and cancer has been investigated. The present study was aimed to clone and produce the Human IL-4 under specific promoter in Escherichia coli and assessed its biological functions. The designed hIL-4 gene construct was artificially synthesized; subsequently, it was sub cloned into the pcDNA3.1 (+) vector in HindIII restriction enzyme site. Recombinant plasmid was transferred and expressed in BL21 cells. The rhIL-4 protein was evaluated by SDS-PAGE and Western blotting. It was purified by Ni-NTA affinity chromatography. The purified protein concentration and also accuracy were determined by ELISA. MTT assay was applied to evaluate the biological activity of rhIL-4 on the Erythroleukemic cell line proliferation. The rhIL-4 gene was successfully cloned and transformed into expression E. coli cells. As a result, a specific band was observed both on the SDS-PAGE and nitrocellulose membrane after Western blotting. The purified protein concentration was equal to 500 pg/ml. The MTT assay indicated that the exposed cells with rhIL-4 were proliferated in a dose dependent manner. The rhIL-4 gene under specific eukaryotic promoter was successfully cloned in the prokaryotic system and the transcription was carried out by T7 RNA polymerase. Therefore, mass production of IL-4 could be a great help in clinical trials and research studies. Additionally, prokaryotic system used in current work was less costly and less time-consuming.
HIGHLIGHTS
- Interleukin-4 (IL-4) is a cytokine that regulates multiple biological functions.
- The recombinant Human IL-4 was cloned and produced under specific promoter in Escherichia coli.
- Through MTT assay, the exposed cells with rhIL-4 were proliferated in a dose dependent manner.
- Cloning
- Human IL-4
- MTT assay
- Prokaryotic system
How to Cite
References
Abbas, A. K., Lichtman, A. H. and S. Pillai, (2014). Cellular and molecular immunology E-book. Eight edition, Elsevier Health Sciences, Philladelphia.
Benczik, M. and S. L. Gaffen, (2004). "The interleukin (IL)-2 family cytokines: survival and proliferation signaling pathways in T lymphocytes." Immunological Investigations, 33(2): 109-142.
Bender, A., Sapp, M., Schuler, G., Steinman, R. M. and N. Bhardwaj, (1996). "Improved methods for the generation of dendritic cells from nonproliferating progenitors in human blood." Journal of Immunological Methods, 196(2): 121-135.
Benedetti, S., Pirola, B., Pollo, B., Magrassi, L., Bruzzone, M. G., Rigamonti, D., Galli, R., Selleri, S., Di Meco, F., De Fraja, C. and A. Vescovi, (2000). "Gene therapy of experimental brain tumors using neural progenitor cells." Nature Medicine, 6(4): 447-450.
Bettelheim, P., Valent, P., Andreeff, M., Tafuri, A., Haimi, J., Gorischek, C., Muhm, M., Sillaber, C.H., Haas, O. and L. Vieder, (1991). "Recombinant human granulocyte-macrophage colony-stimulating factor in combination with standard induction chemotherapy in de novo acute myeloid leukemia." Blood, 77(4): 700-711.
Braun, P. and J. LaBaer, (2003). "High throughput protein production for functional proteomics." Trends in Biotechnology, 21(9): 383-388.
Chomarat, P. and J. Banchereau, (1998). "Interleukin-4 and interleukin-13: their similarities and discrepancies." International Reviews of Immunology, 17(1-4): 1-52.
Corthay, A. (2006). "A three-cell model for activation of naive T helper cells." Scandinavian Journal of Immunology, 64(2): 93-96.
Dauer, M., Obermaier, B., Herten, J., Haerle, C., Pohl, K., Rothenfusser, S., Schnurr, M., Endres, S. and A. Eigler, (2003). "Mature dendritic cells derived from human monocytes within 48 hours: a novel strategy for dendritic cell differentiation from blood precursors." The Journal of Immunology, 170(8): 4069-4076.
Feldmann, M., Brennan, F. M. and R. N. Maini, (1996). "Role of cytokines in rheumatoid arthritis." Annual Review of Immunology, 14: 397-440.
Grone, A. (2002). "Keratinocytes and cytokines." Veterinary Immunology and Immunopathology, 88(1-2): 1-12.
Illera, V. A., Perandones, C. E., Stunz, L. L., Mower, D. A. and R. F. Ashman, (1993). "Apoptosis in splenic B lymphocytes. Regulation by protein kinase C and IL-4." The Journal of Immunology, 151(6): 2965-2973.
Kanhere, A. and M. Bansal, (2005). "Structural properties of promoters: similarities and differences between prokaryotes and eukaryotes." Nucleic Acids Research, 33(10): 3165-3175.
Khan, S., Ullah, M. W., Siddique, R., Nabi, G., Manan, S., Yousaf, M. and H. Hou, (2016). "Role of Recombinant DNA Technology to Improve Life." International Journal of Genomics, 2016: 2405954.
Kitamura, T., Tange, T., Terasawa, T., Chiba, S., Kuwaki, T., Miyagawa, K., Piao, Y. F., Miyazono, K., Urabe, A. and F. Takaku, (1989). "Establishment and characterization of a unique human cell line that proliferates dependently on GM-CSF, IL-3, or erythropoietin." Journal of Cellular Physiology, 140(2): 323-334.
Margolin, K., Aronson, F. R., Sznol, M., Atkins, M. B., Gucalp, R., Fisher, R. I., Sunderland, M., Doroshow, J. H., Ernest, M. L. and J. W. Mier, (1994). "Phase II studies of recombinant human interleukin-4 in advanced renal cancer and malignant melanoma." Journal of Immunotherapy with Emphasis on Tumor Immunology: Official Journal of The Society for Biological Therapy, 15(2): 147-153.
Messeguer, X., Escudero, R., Farré, D., Núñez, O., Martı́nez, J. and M. M. Albà, (2002). "PROMO: detection of known transcription regulatory elements using species-tailored searches." Bioinformatics, 18(2): 333-334.
Morales, J. K., Falanga, Y. T., Depcrynski, A., Fernando, J. and J. J. Ryan, (2010). "Mast cell homeostasis and the JAK-STAT pathway." Genes and Immunity, 11(8): 599-608.
Mosmann, T. R. and R. L. Coffman, (1989). "TH1 and TH2 cells: different patterns of lymphokine secretion lead to different functional properties." Annual Review of Immunology, 7: 145-173.
Mueller, T. D., Zhang, J. L., Sebald, W. and A. Duschl, (2002). "Structure, binding, and antagonists in the IL-4/IL-13 receptor system " Biochimica et Biophysica Acta (BBA)-Molecular Cell Research, 1592(3): 237-250.
Nelms, K., Keegan, A.D., Zamorano, J., Ryan, J. J. and W. E. Paul, (1999). "The IL-4 receptor: signaling mechanisms and biologic functions." Annual Review of Immunology, 17: 701-738.
Overton, T. W. (2014). "Recombinant protein production in bacterial hosts." Drug Discovery Today, 19(5): 590-601.
Paludan, S. R. (1998). "Interleukin-4 and interferon-gamma: the quintessence of a mutual antagonistic relationship." Scandinavian Journal of Immunology, 48(5): 459-468.
Paul, W. E. (1991). "Interleukin-4: a prototypic immunoregulatory lymphokine." Blood, 77(9): 1859-1870.
Raines, R. T., McCormick, M., Van Oosbree, T. R. and R. C. Mierendorf, (2000). "The S.Tag fusion system for protein purification." Methods in Enzymology, 326: 362-376.
Redfield, C., Smith, L. J., Boyd, J., Lawrence, G. M. P., Edwards, R. G., Smith, R. A. and C. M. Dobson, (1991). "Secondary structure and topology of human interleukin 4 in solution." Biochemistry, 30(46): 11029-11035.
Romani, N., Reider, D., Heuer, M., Ebner, S., Kämpgen, E., Eibl, B., Niederwieser, D. and G. Schuler, (1996). "Generation of mature dendritic cells from human blood. An improved method with special regard to clinical applicability." Journal of Immunological Methods, 196(2): 137-151.
Rosano, G. L. and E. A. Ceccarelli, (2014). "Recombinant protein expression in microbial systems." Frontiers in Microbiology, 5: 341.
Rosenberg, H. F., Phipps, S. and P.S. Foster, (2007). "Eosinophil trafficking in allergy and asthma." Journal of Allergy and Clinical Immunology, 119(6): 1303-1310.
Sachdeva, N. and D. Asthana, (2007). "Cytokine quantitation: technologies and applications." Frontiers in Bioscience, 12: 4682-4695.
Seder, R. A., Paul, W. E., Davis, M. M. and B.F. de St Groth, (1992). "The presence of interleukin 4 during in vitro priming determines the lymphokine-producing potential of CD4+ T cells from T cell receptor transgenic mice." Journal of Experimental Medicine, 176(4): 1091-1098.
Shintani, Y., Ito, T., Fields, L., Shiraishi, M., Ichihara, Y., Sato, N., Podaru, M., Kainuma, S., Tanaka, H. and K. Suzuki, (2017). "IL-4 as a Repurposed Biological Drug for Myocardial Infarction through Augmentation of Reparative Cardiac Macrophages: Proof-of-Concept Data in Mice." Scientific Reports, 7(1): 6877.
Struhl, K., Cameron, J. R. and R. W. Davis, (1976). "Functional genetic expression of eukaryotic DNA in Escherichia coli." Proceedings of the National Academy of Sciences, 73(5): 1471-1475.
Terpe, K. (2006). "Overview of bacterial expression systems for heterologous protein production: from molecular and biochemical fundamentals to commercial systems." Applied Microbiology and Biotechnology, 72(2): 211-222.
Van Kimmenade, A., Bond, M.W., Schumacher, J.H., Laquoi, C. and R.A. Kastelein, (1988). "Expression, renaturation and purification of recombinant human interleukin 4 from Escherichia coli." European Journal of Biochemistry, 173(1): 109-114.
Vitetta, E. S., Ohara, J., Myers, C. D., Layton, J. E., Krammer, P. H. and W. E. Paul, (1985). "Serological, biochemical, and functional identity of B cell-stimulatory factor 1 and B cell differentiation factor for IgG1." Journal of Experimental Medicine, 162(5): 1726-1731.
Werwie, N., Sadlek, K., Lundstrom, E., Berrebi, D., Hobbs, G. and L. Vona-Davis, (2017). "The prognostic relevance of inflammatory cytokines and growth factors elaborated from adipose-derived stem cells in breast cancer." Proceedings: AACR Annual Meeting, Abstract 2971.
Whitehead, R. P., Lew, D., Flanigan, R. C., Weiss, G. R., Roy, V., Glode, M. L., Dakhil, S. R. and E. D. Crawford, (2002). "Phase II trial of recombinant human interleukin-4 in patients with advanced renal cell carcinoma: a southwest oncology group study." Journal of Immunotherapy, 25(4): 352-358.
Wong, H. L., Lotze, M. T., Wahl, L. M. and S. M. Wahl, (1992). "Administration of recombinant IL-4 to humans regulates gene expression, phenotype, and function in circulating monocytes." The Journal of Immunology, 148(7): 2118-2125.
Zamorano, J., Rivas, M. D. and G. M. Perez, (2003). "Interleukin-4: A multifunctional cytokine." Immunologia, 22: 215-224.
Zhu, X. and M. Stephens, (2017). "A large-scale genome-wide enrichment analysis identifies new trait-associated genes, pathways and tissues across 31 human phenotypes." bioRxiv, 1:160770.
Zubiaga, A. M., Munoz, E. and B. T. Huber, (1992). "IL-4 and IL-2 selectively rescue Th cell subsets from glucocorticoid-induced apoptosis." The Journal of Immunology, 149(1): 107-112.
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