Saccharomyces cerevisiae secretes and correctly processes human interferon hybrid proteins containing yeast invertase signal peptides.
Synthetic oligonucleotides coding for the yeast invertase secretion signal peptide were fused to the gene for the mature form of human interferon (huIFN-alpha 2). Two plasmids (E3 and F2) were constructed. E3 contained the invertase signal codons in a reading frame with the mature huIFN-alpha 2 gene. F2 had a deletion of the codon for alanine at amino acid residue-5 in the invertase signal and an addition of a methionine codon located between the coding sequences for the invertase signal and mature huIFN-alpha 2. Both hybrid genes were located adjacent to the promoter from the 3-phosphoglycerate kinase gene on the multicopy yeast expression plasmid, YEp1PT. Yeast transformants containing these plasmids produced somewhat more IFN than did the same expression plasmid containing the IFN gene with its human secretion signal sequence. HuIFN-alpha 2, purified from the medium of yeast cells containing E3, was found to be processed at the correct site. The huIFN-alpha 2 made by plasmid F2 was found to be completely processed at the junction between the invertase signal (a variant) and the methionine of methionine-huIFN-alpha 2. These results strongly suggested that the invertase signal (or its variant) attached to huIFN was efficiently recognized by the presumed signal recognition particle and was cleaved by the signal peptidase in the yeast cells. These results also suggested that amino acid changes on the right side of the cleavage site did not necessarily prevent cleavage or secretion.
D000595:Amino Acid Sequence; D004262:DNA Restriction Enzymes; D004926:Escherichia coli / Q000235:genetics; D006026:Glycoside Hydrolases / Q000235:genetics*; D006801:Humans; D007370:Interferon Type I / Q000235:genetics* / Q000378:metabolism; D021382:Protein Sorting Signals / Q000235:genetics*; D011994:Recombinant Proteins / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D043324:beta-Fructofuranosidase
D007370:Interferon Type I; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D004262:DNA Restriction Enzymes; D006026:Glycoside Hydrolases; D043324:beta-Fructofuranosidase