Structural analysis of modified forms of recombinant IFN-beta produced under stress-simulating conditions

The present study focused on the investigation of the chemical stability of recombinant human interferon-beta (rhIFN-beta) tested in vitro by chemical treatments that simulate stress-induced conditions that may occur during h andling, storage or ageing of protein samples, Mild oxidation and/or alkylation of the recombinant protein showed that the four methionines occurring in the interferon displayed different chemical susceptibility in that Met36 and Met117 were fully modified, whereas Met1 s howed only little modification and Met62 was completely resistant, Moreover , incubation of rhIFN-beta under alkaline conditions resulted in the format ion of a covalent dimeric species stabilised by an intermolecular disulphid e bridge involving the free SH group of Cys17 from each polypeptide chain. Analysis of biological activity of the different IFN-beta derivatives showe d that rhIFN-beta fully retains its specific activity following mild oxidat ion treatments whereas reaction with a high concentration of alkylating age nts or incubation under alkaline conditions strongly reduce its specific an tiviral activity.