Author(s): Najim M. Aziz, Mohammed A. AL-Hajaj


DOI: 10.52711/0974-360X.2021.01160   

Address: Najim M. Aziz1*, Mohammed A. AL-Hajaj2
1Department of Marine Biology, Marine Science Center, University of Basrah, Basrah, Iraq.
2Department of Biology, College of Science, University of Basrah, Basrah, Iraq.
*Corresponding Author

Published In:   Volume - 14,      Issue - 12,     Year - 2021

Human interferon-gamma (hIFN-?) is a Glycoprotein pertinence to a distinct group of interferon, called type II interferons, which have an immunological function to respond to antigenic stimuli such as bacteria, viruses, fungi, and any infections by a microorganism. The hIFN-? is produced by natural killer T (NKT) and natural killer (NK) cells during the immune response as part of the innate immune response and by Th1 CD4 and CD8 cytotoxic T lymphocytes (CTL) effector T cells upon the development of antigen-specific immunity. Recombinant hIFN-? has been produced in different expression systems comprising prokaryotic, insect, fungal (yeasts), protozoan, mammalian cells, and plant. In the present study, pET28a plasmid was used in this research for preparation to insert and pYES2 plasmid for cloning and expression of human IFN-? gene in yeast. The trademark of hIFN-? has been produced in Escherichia coli is termed ACTIMMUNE®, but the human interferon- gamma was produced in the prokaryotic expression system is unglycosylated form. This result increases in cost due to increased purification, as well as short-life in the bloodstream, but it is biologically active. This study aimed to use Saccharomyces cerevisiae 4741 strain as a eukaryotic system for expression of hIFN-? cDNA instead of a prokaryotic system in glycosylation patterns as modified translation. But the results are not satisfactory as the produce of yields.

Cite this article:
Najim M. Aziz, Mohammed A. AL-Hajaj. Cloning, Transformation and Expression of Human IFN-γ by Genetic Engineering in Saccharomyces cerevisiae. Research Journal of Pharmacy and Technology. 2021; 14(12):6715-0. doi: 10.52711/0974-360X.2021.01160

Najim M. Aziz, Mohammed A. AL-Hajaj. Cloning, Transformation and Expression of Human IFN-γ by Genetic Engineering in Saccharomyces cerevisiae. Research Journal of Pharmacy and Technology. 2021; 14(12):6715-0. doi: 10.52711/0974-360X.2021.01160   Available on:

1.    Burnett, M.J. and A.C. Burnett, Therapeutic recombinant protein production in plants: Challenges and opportunities. Plants, People, Planet, 2020. 2(2): p. 121-132.
2.    Niharika Mishra, Riyazul Hasan Khan, Bhavna Dwivedi, Sk and Kumar Mishra. Cloning of Gene Coding Glyceraldehyde 3-Phosphate Dehydrogenase. Asian J. Pharm. Tech. 2014; Vol. 4: Issue 4, Oct.-Dec., Pg 184-188.
3.    Nishad Hussain, Jim Eliet Y., Mohammed Muhassinv P., Sherin C.S. Cloning of PA-1L Gene from Pseudomonas aeruginosa in E. coli. Research
J. Engineering and Tech. 4(4): Oct.-Dec., 2013 page 187-190.
4.    T.A., B., <Gene Cloning and DNA Analysis.pdf>. Wiley-Blackwell. 2010. 245-253.
5.    Deepali Anand Sutar, Bhavana. U. Jain, Manish Kondawar. A Review Article on Study of Cloning. Asian J. Res. Pharm. Sci. 2019; 9(2): 148-150.
6.    Kim, H., S.J. Yoo, and H.A. Kang, Yeast synthetic biology for the production of recombinant therapeutic proteins. FEMS yeast research, 2015. 15(1): p. 1-16.
7.    Kraaij, M.D., et al., Human monocytes produce interferon-gamma upon stimulation with LPS. Cytokine, 2014. 67(1): p. 7-12.
8.    Mussakhmetov, A., A. Turgimbayeva, and B. Khassenov, Production Of Recombinant Human Interferon-Gamma In Escherichia Coli. Eurasian Journal of Applied Biotechnology, 2016(2): p. 88-94.
9.    Pestka, S. and S. Baron, Definition and classification of the interferons, in Methods in Enzymology. 1981, Academic Press. p. 3-14.
10.    Tyurin, A.A., Expression of Soluble Active Interferon A in Escherichia coli Periplasm by Fusion with Thermostable Lichenase Using the Domain Insertion Approach. Biochemistry (Moscow), 2017: p. 23.
11.    Ebrahimi, N., et al., Cloning, transformation and expression of human gamma interferon gene in tomato (Lycopersicon esculentum Mill.). Biotechnology & Biotechnological Equipment, 2012. 26(2): p. 2925-2929.
12.    Huang, M., J.-D. Jiang, and Z. Peng, Recent advances in the anti-HCV mechanisms of interferon. Acta Pharmaceutica Sinica B, 2014. 4(4): p. 241- 247.
13.    Markušić, M., et al., Induction of IFN-α Subtypes and Their Antiviral Activity in Mumps Virus Infection. Viral immunology, 2014. 27(10): p. 497-505.
14.    Sadir, R., E. Forest, and H. Lortat-Jacob, The heparan sulfate binding sequence of interferon-γ increased the on rate of the interferon-γ-interferon- γ receptor complex formation. Journal of Biological Chemistry, 1998. 273(18): p. 10919-10925.
15.    Godfrey A, Aisharwya N. M, Babiker B, Gitima D. Role of interleukin -1β in pancreatic islet cells dysfunction and apoptosis in pancreatic islet transplantation. Research J. Pharm. and Tech. 2020; 13(8):3947-3951.
16.    Sareneva, T., et al., Biosynthesis and N‐glycosylation of Human Interferon‐γ: Asn25 and Asn97 Differ Markedly in How Efficiently They are Glycosylated and in Their Oligosaccharide Composition. European Journal of Biochemistry, 1996. 242(2): p. 191-200.
17.    Farrar, M.A. and R.D. Schreiber, The molecular cell biology of interferon- gamma and its receptor. Annual Review of Immunology, 1993. 11(1): p. 571-611.
18.    Trent, J.M., S. Olson, and R.M. Lawn, Chromosomal localization of human leukocyte, fibroblast, and immune interferon genes by means of in situ hybridization. Proceedings of the National Academy of Sciences, 1982. 79(24): p. 7809-7813.
19.    Naylor, S., P. Gray, and P. Lalley, Mouse immune interferon (IFN-γ) gene is on chromosome 10. Somatic cell and molecular genetics, 1984. 10(5): p. 531-534.
20.    Salman, A.H.I.a.M.S., Polymorphism Interferon Gamma Promoter and Receptor among Tuberculosis Patients in Basra Province, South of Iraq. Donnish Journal of Medicine and Medical Sciences 2017. Vol 4(1) pp. 001- 007 January, 2017: p. 8.
21.    Zahra’a, A.A., E.M. Idan, and A.H. Ad'hiah, A Single Nucleotide Polymorphism of Interferon-gamma Gene (IFNG) is not associated with Visceral Leishmaniasis in Iraqi Patients. Current Research in Microbiology and Biotechnology, 2015. 3(5): p. 737-740.
22.    Wang, D., et al., Expression, purification and characterization of human interferon-γ in Pichia pastoris. Molecular Medicine Reports, 2014. 9(2): p. 715-719.
23.    Möller, M., et al., Analysis of eight genes modulating interferon gamma and human genetic susceptibility to tuberculosis: a case-control association study. BMC Infectious Diseases, 2010. 10(1): p. 154.
24.    Rene Devos, H.C., Yoichi Taya, Wim Degrave, Hugo Van Molecular clonmig of human immune interferon cDNA and its expression in eukaryotic cells. IRL Press Umited, 1 Falconberg Court, London WlV 5FG, U.K., 1982. Volume 10 Number 8: p. 15.
25.    Gray, P.W. and D.V. Goeddel, Structure of the human immune interferon gene. Nature, 1982. 298(5877): p. 859.
26.    Bloom, B.R., Interferons and the immune system. Nature, 1980. 284(5757): p. 593-595.
27.    Fleishmann, J.W., Potentiation and inhibition of interferon action: antiviral and antitumor studies. Medical and Pediatric Oncology, 1981. 9(1): p. 89- 92.
28.    Langford, M., et al., Large-scale production and physicochemical characterization of human immune interferon. Infection and Immunity, 1979. 26(1): p. 36-41.
29.    Reddy, S.P., et al., Recombinant interleukin-2 in combination with recombinant interferon-gamma in patients with advanced malignancy: a phase 1 study. Journal of Immunotherapy (Hagerstown, Md.: 1997), 1997. 20(1): p. 79-87.
30.    Gray, P.W., et al., Expression of human immune interferon cDNA in E. coli and monkey cells. Nature, 1982. 295(5849): p. 503.
31.    Gitima Deka, Nagarathna Pkm, Moqbel Ali Moqbel Redhwan, Melvin Mariyam Varghese. IFNγ: Signalling, Epigenetics and Roles in Immunity, Metabolism, Disease and Cancer Immunotherapy. Research J. Pharm. and Tech. 2020; 13(8):4028-4032.
32.    Afshari, E., et al., Cloning, Expression and Purification of Pseudomonas putida ATCC12633 Creatinase. Avicenna Journal of Medical Biotechnology, 2017. 9(4): p. 169.
33.    Zamani, A., K. Oshvandi, and H. Bazmamoun, Lower culture-temperature effects on recombinant IFN-γ production in Chinese Hamster Ovary (CHO) cells. Int. J. Agric. Biol., 2007. 9: p. 378-381.
34.    N. R. Reshma, U. Gayathri. Biological Agents for Delivery of Therapeutic Genes. Research J. Engineering and Tech. 4(4): Oct.-Dec., 2013 page 182- 186.
35.    Pandey, R., A.A. Prabhu, and V.V. Dasu, Purification of recombinant human interferon gamma from fermentation broth using reverse micellar extraction: A process optimization study. Separation Science and Technology, 2017. 53(3): p. 487-495.
36.    Abbaszadeh, S., N. Bakhtiari, and Z. Amini-Bayat, Simple and Effective Purification of Recombinant Peptide Drug, hPTH (1-34), Expressed in E. coli Host. International Journal of Peptide Research and Therapeutics, 2018. 25(2): p. 419-425.
37.    Bhasker. S, Sandeep. G, Ranganath YS. Future of Cancer Therapy-COX-2 Inhibitors: A Review. Research J. Pharm. and Tech.2 (4): Oct.-Dec. 2009; Page617-620.
38.    P. Sai Preethi, M Ramesh Pathy. Recombinant Peroxidase: Production and its Prospective Applications- A Review. Research J. Pharm. and Tech 2018; 11(7): 3186-3196.
39.    Kumar, M., M. Singh, and S.B. Singh, Optimization of conditions for expression of recombinant interferon-γ in E. coli. Molecular Biology Reports, 2014. 41(10): p. 6537-6543.
40.    Mahalik, S., A.K. Sharma, and K.J. Mukherjee, Genome engineering for improved recombinant protein expression in Escherichia coli. Microbial Cell Factories, 2014. 13(1): p. 177.
41.    Saber M. A., O.H., Mohamed M. M., and Romeih M. H., Production of recombinant human interferon gamma by batch fermentation in E. coli system. Arab J. Biotech., 2017. 20: p. 12.
42.    A.Manivannan, K. K. Kumar, S. Varanavasiappan, S. Manimegalai, K. Poornima, B. C. Devrajan, D. Sudhakar, V. Balasubramani. Expression, Purification and Bioassay of Cry55Aa protein against tomato root knot Nematode, Meloidogyne incognita. Res. J. Pharmacognosy and Phytochem. 2020; 12(1):. 19-23.
43    Derynck, R., Expression of the Human Interferon-y cDNA in yeast. 1983. 11: p. 19.
44.    Davoudi, N., et al., Cloning and expression of human IFN-γ in Leishmania tarentolae. World Journal of Microbiology and Biotechnology, 2011. 27(8): p. 1893-1899.
45.    Zamani, A., J.T. Afshari, and M.Y. Alikhani, Molecular Cloning and Expression of Human Gamma Interferon (IFN-g) Full cDNA in Chinese Hamster Ovary (CHO) Cells. Iranian Journal of Immunology, 2006. 3(1): p. 1-8.
46.    Saumya S, Aberami J A, P. Deepa Sankar. Plastid Transformation – A Greener and Cleaner Technique for Overexpression of Proteins. Research J. Pharm. and Tech. 2019; 12(10):5083-5090.
47.    al, B.e., <Designer deletion strains derived from Saccharomyces cerevisiae S288C: a useful set of strains and plasmids for PCR-mediated gene disruption and other applications. Yeast. PubMed: 9483801, 1989. 14: p. 115-132.
48.    Benatuil, L., et al., An improved yeast transformation method for the generation of very large human antibody libraries. Protein Engineering, Design and Selection, 2010. 23(4): p. 155-159.
49.    Wang, Y., et al., Fine-tuning the expression of target genes using a DDI2 promoter gene switch in budding yeast. Sci Rep, 2019. 9(1): p. 12538.
50.    Tang, M., et al., Interferon-gamma-mediated osteoimmunology. Frontiers in immunology, 2018. 9: p. 1508.
51.    Ealick, S.E., et al., Three-dimensional structure of recombinant human interferon-gamma. Science, 1991. 252(5006): p. 698-702.

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