Improved secretion of heterologous proteins by Saccharomyces cerevisiae: effects of promoter substitution in alpha-factor fusions.
The effects of promoter strength on secretion of a heterologous protein, somatomedin-C (SMC), by the yeast Saccharomyces cerevisiae were studied by using the promoters of the MF alpha 1, ACT, and CYC1 genes to control expression of alpha-factor/SMC gene fusions. When a low-copy centromere vector was used to carry the gene fusions in yeast transformants, the greatest secretion was obtained with the MF alpha 1 promoter construction and the least with the CYC1 promoter construction. Unexpectedly, using two types of multicopy vectors, the greatest secretion was obtained with the CYC1 promoter construction and the least with the MF alpha 1 promoter construction. The decrease in secretion by the strongest promoter construction (MF alpha 1 promoter) on multicopy vectors was associated with a decrease in SMC mRNA during growth, a decrease in vector copy number, a decrease in vector stability, and a decrease in transformation frequency. The results demonstrate that, unlike in intracellular expression, promoter strength is not simply related to secretion expression levels. Selection against oversecreting cells during growth may explain the reduced secretion efficiency of strong promoter constructions.
D000199:Actins / Q000235:genetics; D003574:Cytochrome c Group / Q000235:genetics; D005656:Fungal Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D007334:Insulin-Like Growth Factor I / Q000235:genetics* / Q000378:metabolism; D000072235:Mating Factor; D010455:Peptides / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D013002:Somatomedins / Q000235:genetics*; D014170:Transformation, Genetic
D000199:Actins; D003574:Cytochrome c Group; D005656:Fungal Proteins; D010455:Peptides; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D013002:Somatomedins; D000072235:Mating Factor; D007334:Insulin-Like Growth Factor I