Vectors for the inducible overexpression of glutathione S-transferase fusion proteins in yeast.
A rapid and convenient method of protein purification involves creating a fusion protein with glutathione S-transferase (GST) (Smith and Johnson, Gene 67, 31-40, 1988). In this report, we describe two vectors for the conditional expression of GST fusions in Saccharomyces cerevisiae. The parent plasmid is based on a high-copy, galactose-inducible shuttle vector previously described (Baldari et al., EMBO J. 6, 229-243, 1987). We have demonstrated the use of this system by creating fusions between GST and the yeast RAS2 gene. GST-Ras2 fusion proteins undergo the post-translational modifications required for Ras2p to become membrane localized. These vectors provide a useful system for the expression and purification of eukaryotic proteins requiring post-translational modification.
Mitchell
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Department of Biochemistry, University of Iowa, Iowa City 52242.
Marshall
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Deschenes
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RJ
D001483:Base Sequence; D005656:Fungal Proteins / Q000235:genetics; D005690:Galactose; D015870:Gene Expression / Q000235:genetics; D005800:Genes, Fungal / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D005982:Glutathione Transferase / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D018631:ras Proteins
D005656:Fungal Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D005982:Glutathione Transferase; C489015:RAS2 protein, S cerevisiae; D018631:ras Proteins; D005690:Galactose