High yield expression, characterization, and biological activity of IFNα2-Tα1 fusion protein.

The use of interferon alpha-2 in combination with thymosin alpha-1 shows higher anti-cancer effect in comparison when both are used individually because of their synergistic effects. In this study we produced an important human interferon alpha-2-thymosin alpha-1 (IFN alpha 2-T alpha 1) fusion protein with probable pharmaceutical properties coupled to its high-level expression, characterization, and study of its biological activity. The IFN alpha 2-T alpha 1 fusion gene was constructed by over-lap extension PCR and expressed in Escherichia coli expression system. The expression of IFN alpha 2-T alpha 1 fusion protein was optimized to higher level and its maximum expression was obtained in modified terrific broth medium when lactose was used as inducer. The fusion protein was refolded into its native biologically active form with maximum yield of 83.14% followed by purification with similar to 98% purity and 69% final yield. A band of purified IFN alpha 2-T alpha 1 fusion protein equal to similar to 23 kDa was observed on 12 % SDS-PAGE gel. The integrity of IFN alpha 2-T alpha 1 fusion protein was confirmed by western blot analysis and secondary structure was assessed by CD spectroscopy. When IFN alpha 2-T alpha 1 fusion protein was subjected to its biological activity analysis it was observed that it exhibits both IFN alpha 2 & T alpha 1 activities as well as significantly higher anticancer activity as compared to IFN alpha-2 alone.