Secretion of Escherichia coli beta-galactosidase in Saccharomyces cerevisiae using the signal sequence from the glucoamylase-encoding STA2 gene.
The budding yeast Saccharomyces cerevisiae is a safe and widely used host for the production of recombinant DNA-derived proteins. We have used the signal sequence from the S. diastaticus STA2 gene, encoding glucoamylase II, to secrete Escherichia coli beta-galactosidase, encoded by the lacZ gene. In frame STA2/lacZ gene fusions have been constructed and expressed in S. cerevisiae under the control of either the STA2 or the galactose inducible GAL1-10 upstream promoters. Fairly high amounts of the enzyme (up to 76% of total activity, depending on the growth conditions) are secreted in the periplasmic space. Adding yeast extract and peptone to the growth medium results in a dramatic increase in both synthesis and secretion of beta-galactosidase.
Vanoni
M
M
Dipartimento di Fisiologia e Biochimica Generali, Università degli Studi di Milano, Italy.
Porro
D
D
Martegani
E
E
Alberghina
L
L
D000595:Amino Acid Sequence; D001483:Base Sequence; D004274:DNA, Recombinant / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D005696:Galactosidases / Q000235:genetics*; D015870:Gene Expression; D005798:Genes, Bacterial; D005087:Glucan 1,4-alpha-Glucosidase / Q000235:genetics*; D015151:Immunoblotting; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics*
D004274:DNA, Recombinant; D021382:Protein Sorting Signals; D005696:Galactosidases; D001616:beta-Galactosidase; D005087:Glucan 1,4-alpha-Glucosidase