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

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Abstract
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.