title: The expression in Escherichia coli of a synthetic gene coding for the precursor of papain is prevented by its own putative signal sequence.
abstract: A 1048-bp gene coding for prepropapain was assembled from chemically synthesized oligodeoxyribonucleotides and cloned into a variety of Escherichia coli expression plasmids. We observed loss of plasmid when the preproP gene was expressed in E. coli either as the native precursor or fused at the C terminus of the first 592 amino acids (aa) of beta-galactosidase (beta Gal). Deletion of the putative 26-aa signal peptide (pre-region) increased plasmid stability. The level of maintenance for the different plasmid constructs correlated with the level of expression detected by immunoblotting. Constitutive expression of the beta Gal-propapain fusion generated insoluble granules in a protease-deficient E. coli host. The fusion protein was easily purified to near homogeneity by differential solubilization of the granules.
authors.affiliation: Biotechnology Research Institute, National Research Council of Canada, Montréal, Québec.
authors.forename: T
authors.initials: T
authors.lastname: Vernet
authors.forename: D C
authors.initials: DC
authors.lastname: Tessier
authors.forename: F
authors.initials: F
authors.lastname: Laliberté
authors.forename: D
authors.initials: D
authors.lastname: Dignard
authors.forename: D Y
authors.initials: DY
authors.lastname: Thomas
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D004247:DNA; D004926:Escherichia coli / Q000235:genetics*; D005786:Gene Expression Regulation; D005813:Genes, Synthetic; D008969:Molecular Sequence Data; D009154:Mutation; D010206:Papain / Q000096:biosynthesis / Q000235:genetics*; D010957:Plasmids; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics*; D021382:Protein Sorting Signals / Q000235:genetics / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D011498:Protein Precursors; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D004247:DNA; D010206:Papain