Alpha-factor leader sequence-directed transport of Escherichia coli beta-galactosidase in the secretory pathway of Saccharomyces cerevisiae

Mol Gen Genet. 1989 Aug;218(2):240-8. doi: 10.1007/BF00331274.

Abstract

The construction of two fused genes is described. One involves the in-frame fusion of the yeast prepro-alpha-factor coding sequence, and the Escherichia coli lac Z gene. The second gene fusion utilizes a 103 bp yeast invertase NH2-terminal coding sequence at the fusion junction of the hybrid gene described above. The gene fusions, under the control of the alpha-factor promoter, expressed active beta-galactosidase in alpha haploid yeast cells. The activity could be regulated in a temperature-sensitive sir3 mutant. The incorporation of the invertase coding sequence at the MF alpha 1-lacZ fusion junction provided significantly higher levels of beta-galactosidase activity. A substantial quantity of the hybrid proteins generated from the gene fusions was primarily localized in the intracellular membranes of yeast cells, while a processed form could be secreted into the periplasm.

MeSH terms

  • Blotting, Northern
  • Blotting, Western
  • Cell Fractionation
  • Cloning, Molecular
  • Culture Media
  • Escherichia coli / enzymology
  • Escherichia coli / genetics*
  • Galactosidases / genetics*
  • Gene Expression Regulation
  • Mating Factor
  • Peptides / genetics*
  • Precipitin Tests
  • Recombinant Fusion Proteins / analysis
  • Saccharomyces cerevisiae / genetics*
  • Subcellular Fractions / analysis
  • beta-Galactosidase / genetics*

Substances

  • Culture Media
  • Peptides
  • Recombinant Fusion Proteins
  • Mating Factor
  • Galactosidases
  • beta-Galactosidase