Abstract
This chapter discusses the purification of recombinant human fibroblast interferon produced in Escherichia coli . Many interferons have been expressed in Escherichia coli . Their use, however, requires purification to homogeneity so that traces of E. coli–– endotoxin and other contaminants––are undetectable or at acceptable levels. The purification of human interferon beta (Hu-IFN-β) expressed in E. coli is described. The following procedure has been used to isolate recombinant human fibroblast interferon from up to 3 kg of Escherichia coli paste. The separation of correctly refolded IFN-α2 from contaminating conformational variants is not a trivial problem. These variants have similar properties to the native molecule, and purification strategies are complicated by the tendency of IFN-α2 to form oligomers transiently in concentrated solution (>1 mg/ml). Conformational variants that participate in this association are consequently almost impossible to completely separate from the “native” conformer. The purification approach incorporates separation techniques selected for their compatibility with buffer systems favoring the monomeric form of the protein: high ionic strength and low pH.
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