Mutations That Allow Disulfide Bond Formation in the Cytoplasm of Escherichia coli

• Published in: Science (American Association for the Advancement of Science) Vol. 262; no. 5140; pp. 1744 - 1747
• Main Authors: Derman, Alan I., Prinz, William A., Belin, Dominique, Beckwith, Jon
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for the Advancement of Science 10.12.1993; American Association for the Advancement of Science
• Subjects: Alkaline Phosphatase - chemistry; Alkaline Phosphatase - metabolism; Amino acids; Bacteriology; Biological and medical sciences; Cysteine - metabolism; Cytoplasm; Cytoplasm - enzymology; Disulfides; Disulfides - metabolism; Escherichia coli; Escherichia coli - enzymology; Escherichia coli - genetics; Escherichia coli - growth & development; Fundamental and applied biological sciences. Psychology; Genes; Genes, Bacterial; Genetic aspects; Genetic mutation; Ice; Metabolism. Enzymes; Microbial mutation; Microbiology; Mutation; Oxidation-Reduction; Plasmids; Protein Folding; Protein Sorting Signals; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Spheroplasts; Thioredoxin; Thioredoxin-Disulfide Reductase - genetics; Thioredoxin-Disulfide Reductase - metabolism; Urokinase-Type Plasminogen Activator - chemistry; Urokinase-Type Plasminogen Activator - metabolism; Proteins; Enzymatic activity; Structure activity relation; Thioredoxin reductase (NADPH); Enzyme; Disulfide bond; Escherichia coli; Bacteria; Oxidoreductases; Mutation; Cytoplasm; Enterobacteriaceae
• ISSN: 0036-8075; 1095-9203
• DOI: 10.1126/science.8259521

Disulfide bonds are rarely found in cytoplasmic proteins. Mutations were selected for in Escherichia coli that allow disulfide bond formation in the cytoplasm. In the presence of these mutations, export-defective versions of alkaline phosphatase and mouse urokinase were able to fold into their enzymatically active conformations in the cytoplasm because their disulfide bonds were formed. The mutations were mapped to the gene for thioredoxin reductase and diminish or eliminate the activity of this enzyme. Thioredoxin itself was found to be unnecessary for this disulfide bond formation. Thioredoxin reductase, but not thioredoxin, is thus implicated in keeping cysteines reduced in cytoplasmic proteins.