Mutational Analysis of the Superantigen Staphylococcal Exfoliative Toxin A (ETA)

Abstract

Exfoliative toxin A (ETA) is known to be a causative agent of staphylococcal scalded skin syndrome (SSSS). Although relatively little is known about exactly how the exfoliative toxins (ETs) cause SSSS, much has been discovered recently that may help elucidate the mechanism(s) by which ETA exhibits activities such as lymphocyte mitogenicity and epidermolytic activity. Here, we have shown that highly purified ETA does have T lymphocyte mitogenic activity in that wild-type ETA induced T cell proliferation whereas several single amino acid mutants lacked significant activity. Neither wild-type ETA nor any single amino acid mutants were proteolytic for a casein substrate, yet esterase activity was detected in wild-type ETA and several mutants, but eliminated in other mutants. A mutation in aa 164 (Asp to Ala) showed a 9-fold increase in esterase activity as well. Finally, we correlated esterase activity with epidermolytic activity. All mutants that lost esterase activity also lost epidermolytic activity. Conversely, mutants that retained esterase activity also retained exfoliative activity, implicating serine protease or serine protease-like activity in the causation of SSSS. Moreover, the mutants that displayed markedly reduced T cell superantigenic activity retained their epidermolytic activity (although some of these mutants required higher doses of toxin to cause disease), which suggests an ancillary role for this activity in SSSS causation.

Staphylococcus aureus Isolated in Cases of Impetigo Produces Both Epidermolysin A or B and LukE-LukD in 78% of 131 Retrospective and Prospective Cases

“…However, detection of the toxin in these bullae was never undertaken. Finally, Rago et al (38) further investigated the topic and reported that the mitogenic activity associated with epidermolysins was not responsible for the lesions observed in the experimental model, compared to the esterase activity. Schlievert et al identified a proteolytic activity that could be exerted on ␣-and ␤-melanoma-stimulating hormones (43).…”

“…This would corroborate epidemiological observations and explain the purulent exudates observed in impetigo bullae or erosive manifestations. There is a lack of completely satisfying experimental animal models for epidermolysin and LukE-LukD and possibly also for the mitogenic activity of epidermolysins tested in the lymphocytes of rabbits (38).…”

“…However, a scarlatiniform erythroderma may be observed in some cases that are not associated with TSST-1 or enterotoxins (16,23). The mode of action of epidermolysins remained controversial until recently (23), but at present a proteolytic activity is presumed (1,38). However, this does not explain the presence of aqueous or purulent exudates in the bullae.…”

Biochemical and Biological Properties of Staphylococcal Enterotoxin K

“…The vector plasmid contains TT overhangs that allow for cloning of PCR products directly without restriction digestion. Subsequently, the gene was excised from this vector with EcoRI, using restriction sites on either side of the insertion site in the vector, and ligated with EcoRI-digested plasmid vector pCE104 (44). The resultant plasmid was transformed into Escherichia coli XL-1 Blue (Stratagene, La Jolla, Calif.), and a DNA insert with a restriction fragment of the proper length was verified by plasmid purification with the Qiagen Spin Miniprep kit (Qiagen, Valencia, Calif.), digestion with EcoRI, and separation in 1% agarose gels.…”

Staphylococcal scalded skin syndrome in a neonate

“…3 These toxins induce proteolysis by trypsin like serine proteases. 4 ETA exhibits superantigen activities.…”