Our research involves exploration of the biology of the staphylococcal enterotoxins (SEs). One project in the lab is identification and characterization of previously unidentified SEs. Two SEs, designated SEG and SEI, have been identified by hybridization with a sec gene probe. The sequence of each has been determined, indicating that SEG is more closely related to SEB and SEC while SEI is more closely related to SEA, SED and SEE. Culture supernatants from a strains containing cloned seg or sei genes stimulate proliferation of murine splenocytes. Culture supernatants from these strains will also be tested for ability to provoke an emetic response in rhesus monkeys. A histidine-tagged SEG construct has been made for the production of polyclonal antibodies against SEG for use in detection of SEG in culture supernatants. Wild-type SEG is also being purified from the seg gene clone for use in MHC class II binding assays. Similiarly, histidine-tagged SEI is being constructed for similar studies. The second major project in the lab is structure function analysis of SEA. The general strategy in these studies has been the construction of SEAs with amino acid substitutions or deletions. These mutant SEAs are tested for emetic and superantigen activity. Our studies have identified one site in SEA in which substitutions affect emetic activity but not ability to stimulate murine splenocytes; other substitutions which affect superantigen but not emetic activity have also been identified. Further studies using in vitro stimulation of rhesus monkey peripheral blood leukocytes with SEA and mutant SEAs are being performed to examine the superantigenic activation of rhesus T-cells at the molecular level. Key words staphylococcal enterotoxins, SEA, SEG, SEI, primate emetic assay, superantigen
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