Expression of a functional human tumor necrosis factor-α (hTNF-α) in Yeast Saccharomyces cerevisiae

The recombinant soluble human tumor necrosis factor-alpha (hTNF-α) was expressed in a yeast Saccharomyces cerevisiae and its cytotoxicity was evaluated. A cDNA encoding hTNF-α was placed under the control of two different promoters: a glyceraldehyde-3-phosphate dehydrogenase (GPD) promoter and a yeast hybrid ADH2-GPD promoter, consisting of alcohol dehydrogenase II (ADH2) and the GPD promoter. A Northern blot analysis revealed that, although variation in the expression level of hTNF-α existed among transformants, the higher expression was obtained with the GPD promoter. Expressed hTNF-α protein (rhTNF-α) was successfully secreted into the culture medium, producing 2.5 mg per liter of culture filtrate, with no changes in cell growth. The bioassay for observing the cytotoxicity to the murine L929 fibroblast cell line, with serial dilution of rhTNF-α, indicated that the secreted rhTNF-α was bioactive and its doseresponse was improved eight to ten times over that of the E. coli-derived rhTNF-α.