Efficient synthesis of secreted murine interleukin-2 by Saccharomyces cerevisiae: influence of 3'-untranslated regions and codon usage.
Several expression vectors were compared which directed the synthesis of secreted murine interleukin-2 (mIL2) in the culture medium of Saccharomyces cerevisiae. We used the prepro-sequence of the alpha 1 mating-factor precursor as a secretion signal in S. cerevisiae in combination with different promoters. The yield of mature mIL2 was significantly improved by deleting the major part of the 3'-untranslated region (UTR). In Northern-blotting experiments we showed that a destabilizing sequence present in the 3' UTR might be responsible for rapid degradation of the mIL2 mRNA. The highest expression (about 10 micrograms/ml) was obtained under control of the GAL1 promoter in an S. cerevisiae strain where the regulatory GAL4 gene was overexpressed. No difference in expression level was observed in a construct wherein twelve consecutive codons were replaced by optimal codons for S. cerevisiae.
Demolder
J
J
Laboratory of Molecular Biology, State University, Gent, Belgium.
Fiers
W
W
Contreras
R
R
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D003062:Codon / Q000235:genetics*; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000235:genetics; D015966:Gene Expression Regulation, Fungal / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D007376:Interleukin-2 / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D000072235:Mating Factor; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011498:Protein Precursors / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors / Q000235:genetics
D003062:Codon; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D007376:Interleukin-2; D010455:Peptides; D011498:Protein Precursors; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D000072235:Mating Factor