Targeted mutations in Val101 and Arg27 interferon beta protein increase its transcriptional and translational activities. (February 2016)

• Record Type: Journal Article
• Authors: Kay, Maryam; Hasan-Abad, Amin Moradi; Hojati, Zohreh; Korbekandi, Hasan
• Is Part Of: Cytokine. Volume 78(2016)
• Page Start: 1
• Page End: 6
• Publication Date: 2016-02
• Subjects: ARE elements -- Codon 101 -- Interferon beta -- Real-time PCR; Cytokines -- Periodicals; 571.844
• DOI: 10.1016/j.cyto.2015.11.019
• Languages: English
• ISSNs: 1043-4666

Abstract

Highlights: Enhancement of Interferon beta production by site directed mutagenesis. Two recombinant constructs including IFNβ101, IFNβ101+27 were used. Elevated RNA and protein expression was seen with IFNβ101 and IFNβ101+27 . Abstract: Interferon β (IFNβ) is the most prescribed drug that has been used frequently for the treatment of multiple sclerosis (MS) patients. The aim of this study is to improve the production of IFNβ by induction of site directed mutagenesis. Accordingly, recombinant constructs were designed in order to enhance the expression of IFNβ mRNA and protein. The recombinant plasmids were transfected to the CHO cell line, following RNA extractions and cDNA synthesis. The effects of recombinant constructs were analyzed by real time PCR, ELISA and MTT assay. Transfected samples with either IFNβ 101 or IFNβ 101+27 have shown 11.55 and 2.26 fold elevation and over-expression compare to the wild type construct respectively. Our data also indicated that the IFNβ101 and IFNβ101+27 constructs increase IFNβ protein expression more than 2.2 and 4.5 fold, respectively compared to the control group. It could be concluded that the substitution of Phe in the codon 101 position, which may increase the binding activity of IFNβ with its receptors and introduction of an additional N glycosylation site (Asn-X-Thr) in the position 27 of IFNβ protein may cause such an effect. The proliferative activity of transfected cells by a recombinant IFNβ101 decreases in comparison to the wild type, although it was not statistically significant. Over-expression of IFNβ in such a level is promising not only for the patients but also for the pharmaceutical industries.