Formation of disulfide bonded dimer of mutated heat-labile enterotoxin in vivo

One of the two cysteines in the B subunit of heatlabile enterotoxin has been changed to a serine by site-directed mutagenesis so that the internal disulfide bond cannot form. The mutant protein, like the wild-type protein synthesised in the presence of the reducing agent dithiothreitol, does not for...

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Bibliographic Details
Published in:FEBS letters 1996-02, Vol.381 (1), p.63-66
Main Authors: Hedges, Peter A., Hardy, Simon J.S.
Format: Article
Language:English
Subjects:
Bacterial Toxins - biosynthesis
Bacterial Toxins - chemistry
Bacterial Toxins - isolation & purification
Cell Membrane - metabolism
Disulfide bond
Disulfides
Dithiothreitol - pharmacology
DTT, dithiothreitol
Electrophoresis, Polyacrylamide Gel
Enterotoxins - biosynthesis
Enterotoxins - chemistry
Enterotoxins - isolation & purification
Escherichia coli - metabolism
Escherichia coli Proteins
Genes, Bacterial
Heat-labile enterotoxin
IPTG, isopropylthiogalactoside
Kinetics
Macromolecular Substances
Membrane association
Molecular Weight
Mutagenesis
Recombinant Proteins - biosynthesis
Recombinant Proteins - chemistry
Recombinant Proteins - isolation & purification
Spheroplasts - metabolism
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Summary:One of the two cysteines in the B subunit of heatlabile enterotoxin has been changed to a serine by site-directed mutagenesis so that the internal disulfide bond cannot form. The mutant protein, like the wild-type protein synthesised in the presence of the reducing agent dithiothreitol, does not form pentamers in the periplasm but binds to available membranes. Binding to membranes is disrupted by chaotropic agents but not by salt. More than half the molecules of mutant protein form disulfide-bonded dimers when exported to the periplasm but no dimer is detected when the protein is exported to the medium by spheroplasts.
ISSN:0014-5793
1873-3468
DOI:10.1016/0014-5793(96)00082-8