Development of the chromatographic medium for the affinity isolation of the recombinant hIFN-β1b based on immobilized single-chain antibodies

Abstract

Aim. The development of a laboratory method for the production of immunoaffi nity chromatography medium for the purifi cation of the recombinant human IFN-β1b. Methods. A gene of the chimeric protein ScFv bINF -CBD was constructed using the DNA sequences encoding ScFv specifi c to hIFN-β1b and the cellulose-binding domain (CBD) of Clostridium thermocellum. The developed chimeric protein was expressed in Escherichia coli cells. The target protein was obtained in soluble, functionally active form by its renaturation from the bacterial inclusion bodies in vitro. The ScFv bINF -CBD was immobilized on a chitin carrier. Results. The introduction of CBD by gene engineering techniques enabled the oriented non-covalent immobilization of ScFv bINF -CBD on chromatographic matrix. The developed immunoaffi nity medium allowed isolating the rhIFN-β1b from the complex mixture, after its renaturation from the inclusion bodies, with more than 89 % purity. Conclusion. The designed immunoaffi nity medium provides isolating the rhIFN-β1b from the complex protein mixtures. K e y w o r d s: Interferon-β1b, single-chain antibodies, immunoaffi nity purifi cation, cellulose-binding domain.