29] Purification of recombinant human immune interferon

Abstract

Publisher Summary This chapter discusses the purification of recombinant human immune interferon. Full length complementary DNA (cDNA) coding for mature human immune interferon (IFN-γ) has been isolated, and the coding sequence for mature IFN-γ has been expressed in Escherichia coli, yeast, and monkey cells. The purification as well as the structural characterization of the bacterial product is described. The purification of IFN-γ can be carried out simply, efficiently, and rapidly, with the use of silica gel and monoclonal antibodies to the COOH-terminal peptide of IFN-γ. IFN-γ appears to be degraded during the purification process. The degradation can be prevented with the use of guanidine hydrochloride in the initial step of purification, and homogeneous intact molecules could be obtained (18K IFN-7). By contrast, the sonication of frozen cells in the absence of guanidine yielded mainly the truncated product (15K IFN-γ, where 15 COOH-terminal amino acid residues are lost). Amino acid compositions and NH2-terminal sequences are consistent with that expected from the DNA sequence. The specific activities of the 18K and 15K IFN-γ are the same (107 units/mg) and comparable to that of the homogeneous natural human immune interferon.