Production and purification of recombinant human interleukin‐6 secreted by the yeast Saccharomyces cerevisiae

The coding region of the human interleukin‐6 (hIL6) gene was fused to the prepro secretion signal of the α‐mating factor gene in several yeast host strains. It was found that the KEX‐2 protease was unable to cleave the prepro‐Lys‐Arg‐Pro‐IL6 sequence, but that unspecific cleavage of the precursor protein had occurred. The prepro‐Lys‐Arg‐Ala‐Pro‐IL6 sequence, however, was correctly recognized and cleaved by the KEX‐2 protease, and IL6 was efficiently secreted into the culture medium. The N‐terminal Ala‐Pro peptide was removed during processing by wild‐type yeast strains, but was retained in a ste13 mutant. IL6 as well as the aberrant proteins were not glycosylated. The transformed cells could secrete up to 30 μg/ml IL6. The protein was purified from the medium to homogeneity by ion‐exchange chromatography and gel filtration, and had a specific activity of about 2 × 108 IU/mg in a proliferation assay.