Cloning of the gene coding for staphylococcal exfoliative toxin A and its expression in Escherichia coli

Sakurai, S

doi:10.1016/0378-1097(87)90499-x

Cited by 7 publications

(12 citation statements)

References 5 publications

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“…The association between SSSS and S. aureus was suspected soon after the condition was described over a hundred years ago [29] but it was not until 1970 that Melish and Glasgow were able to show that an exoprotein of S. aureus was responsible for the exfoliation in newborn mice [2]. This exotoxin was soon isolated, sequenced, characterised and cloned into other bacteria such as Escherichia coli [30–36]. S. aureus produces at least four antigenically distinct toxin serotypes of which two, exfoliative toxins A (ETA) and B (ETB), are responsible for most human cases of SSSS [4].…”

Section: The Toxins As Enzymesmentioning

confidence: 99%

Understanding the mechanism of action of the exfoliative toxins ofStaphylococcus aureus

Ladhani¹

2003

FEMS Immunology &Amp; Medical Microbiology

108

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The exfoliative toxins of Staphylococcus aureus are responsible for the staphylococcal scalded skin syndrome, a blistering skin disorder that particularly affects infants and young children, as well as adults with underlying disease. Their three-dimensional structure is similar to other glutamate-specific trypsin-like serine proteases with two substrate-binding domains and a serine-histidine-aspartate catalytic triad that forms the active site. However, unlike other serine proteases, the exfoliative toxins possess a highly charged N-terminal alpha-helix and a unique orientation of a critical peptide bond, which blocks the active site of the toxins so that, in their native state, they do not possess any significant enzymatic activity. The target for the toxins has recently been identified as desmoglein-1, a desmosomal glycoprotein which plays an important role in maintaining cell-to-cell adhesion in the superficial epidermis. It is speculated that binding of the N-terminal alpha-helix to desmoglein-1 results in a conformation change that opens the active site of the toxin to cleave the extracellular domain of desmoglein-1 between the third and fourth domains, resulting in disruption of intercellular adhesion and formation of superficial blisters. Elucidating the mechanism of action of the toxins and identifying desmoglein-1 as their specific epidermal substrate has not only given us an insight into the pathogenesis of the staphylococcal scalded skin syndrome, but also provided us with useful information on normal skin physiology and the pathogenesis of other toxin-mediated diseases. It is hoped that this knowledge will lead to development of rapid screening and diagnostic tests, and new antitoxin strategies for the treatment and prevention of the staphylococcal scalded skin syndrome in the near future.

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Section: The Toxins As Enzymesmentioning

confidence: 99%

Understanding the mechanism of action of the exfoliative toxins ofStaphylococcus aureus

Ladhani¹

2003

FEMS Immunology &Amp; Medical Microbiology

108

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“…ETA is encoded by a bacterial chromosome, whereas ETB is encoded by a plasmid (20). Both genes have been sequenced (7,13), and the ETA gene has been cloned in Escherichia coli (15).…”

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confidence: 99%

Purification of Protease from a Mixture of Exfoliative Toxin and Newborn-Mouse Epidermis

2000

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Although the role of exfoliative toxin in staphylococcal scalded-skin syndrome has been suggested to be that of a serine protease, it has not been demonstrated to show proteolytic activity. Our purpose was to purify a proteolytic enzyme from a mixture of exfoliative toxin and newborn-mouse epidermis. We used gel filtration and ion-exchange and hydroxyapatite chromatography with a high-pressure liquid chromatography system. A casein-hydrolyzing enzyme was isolated from the mixture. The molecular mass of the enzyme was confirmed to be 20 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Subcutaneous injection of the purified enzyme into newborn mice reproduced the epidermal splitting that is seen in staphylococcal scalded-skin syndrome. These results suggest that exfoliative toxin does not work as a protease itself but that some reaction between exfoliative toxin and an epidermal component(s) first produces a protease, after which epidermal splitting occurs.

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“…We have reported two serotypes, exfoliative toxin A (ETA) and B (ETB) (7)(8)(9), with equal biological activity. Both the eta and etb genes have been cloned and sequenced (12,16,17,19,20). SSSS occurs mostly in newborns, infants, and young children.…”

Section: Abstract : Staphylococcal Exfoliative Toxin B Staphylococcmentioning

confidence: 99%

New Evidence That the Tyr‐157 and Tyr‐159 Residues of Staphylococcal Exfoliative Toxin B Are Essential for Its Toxicity

Sakurai

Suzuki

Saito

et al. 1998

Microbiology and Immunology

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To determine the active site of exfoliative toxin B (sETB) of Staphylococcus aureus, the etb gene was cloned from an S. aureus SU strain obtained from a patient with impetigo. We prepared a frame shift mutant protein from a recombinant plasmid with a BglII linker inserted into the Tyr-155 codon of the ETB gene (pETB/BgIIIL). The recombinant mutant protein (ETB/BgIIIL) obtained from Escherichia coli containing pETB/BgIIIL showed no toxicity in neonatal mice and no agglutination activity. The 20-kDa ETB/BgIIIL contained 185 amino acid residues. Site-directed mutagenesis was used to introduce mutations at either Tyr-155, Tyr-157, Tyr-159, or Tyr-162. Substitution of any of the Tyr residues decreased exfoliative activity compared with that of native sETB (4,000 EU/ml). Substitution of Tyr-155 with a Phe (ETB/Y155) decreased activity 5-fold (800 EU/ml). Substitution of Tyr-157 with Leu (ETB/Y157) decreased activity 80-fold (50 EU/ml) and decreased agglutination titer 5-fold compared with that of native sETB (400,000). Substitution of Tyr-159 with Leu (ETB/Y159)decreased activity 4-fold (1,000 EU/ml). When both Tyr-157 and Tyr-159 were mutated (ETB/Y157-159), both toxicity and antigenicity were completely lost. On an immunodiffusion test, ETB/Y157 showed a faint precipitation line, but ETB/BgIIIL and ETB/Y157-159 had no activity, showing that the Tyr-157 and Tyr-159 residues are essential for the toxicity and antigenicity of ETB.

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Cloning of the gene coding for staphylococcal exfoliative toxin A and its expression in Escherichia coli

Cited by 7 publications

References 5 publications

Understanding the mechanism of action of the exfoliative toxins ofStaphylococcus aureus

Understanding the mechanism of action of the exfoliative toxins ofStaphylococcus aureus

Purification of Protease from a Mixture of Exfoliative Toxin and Newborn-Mouse Epidermis

New Evidence That the Tyr‐157 and Tyr‐159 Residues of Staphylococcal Exfoliative Toxin B Are Essential for Its Toxicity

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