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Molecular and Biological Characterization of a Prepared Recombinant Human Interferon Alpha 2b Isoform

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Abstract

Recombinant human interferon alpha2b (rhIFN-α2b) protein is FDA approved for treatment of many tumors and viral diseases. A rhIFN-α2b isoform has been produced and purified from the refolding reaction using high-resolution anion ion exchange chromatography. This isoform has a proper MW (19 kDa) and high purity and homogeneity. The conservation of native linear and conformational epitopes in this isoform was immunologically confirmed by Western blot and ELISA. Mass spectrometry assessment of its intact mass showed average mass (19,337 Da) equivalent to that of the expressed rhIFN-α2b protein without any chemical modification and without the first methionine. Peptide mapping of rhIFN-α2b through tryptic digestion of reductive/alkylated protein using urea as a denaturing agent gave the best pattern. The rhIFN-α2b had a high specific antiviral activity (2.5 × 108 ± 1.1 × 108IU/mg protein). In vivo clearance study of rhIFN-α2b in female SD rats (500 μg/kg, intramuscularly) revealed rapid clearance (elimination half-life 0.54 h with a maximum plasma concentration of 33,792 pg/ml) compared with the commercial rhIFN-α2 (elimination half-life 0.75–0.96 h). In conclusion, the prepared rhIFN-α2b isoform has high purity, homogeneity, native like chemical and structural composition, high antiviral activity, and proper biological stability, which reduce its immunogenicity and raise its therapeutic efficiency.

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Acknowledgements

Thanks are due to Prof. Mohamed Salah Eldin Mohamed (Parasitology Department, Theodor Bilharz Research Institute) for helping in experimental animal part.

Funding

This work was supported by the STDF project Grant No. 1454.

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Theodor Bilharz Research Institute, P.O. Box 30, Giza, Egypt

    Rasha Shaaban, Safia Samir & Ehab El-Dabaa

  2. Faculty of Science, Department of Zoology, University of Ain Shams, Abbassia, Cairo, 11566, Egypt

    Wael M. El-Sayed

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  1. Rasha Shaaban
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  2. Wael M. El-Sayed
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  4. Ehab El-Dabaa
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Correspondence to Wael M. El-Sayed or Ehab El-Dabaa.

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Shaaban, R., El-Sayed, W.M., Samir, S. et al. Molecular and Biological Characterization of a Prepared Recombinant Human Interferon Alpha 2b Isoform. Appl Biochem Biotechnol 188, 72–86 (2019). https://doi.org/10.1007/s12010-018-2908-y

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  • DOI: https://doi.org/10.1007/s12010-018-2908-y

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