BIOCHEMICAL AND MUTATIONAL ANALYSIS OF THE HISTIDINE-RESIDUES OF STAPHYLOCOCCAL-ENTEROTOXIN-A

The goal of this study was to examine the role of histidine residues i n the biological activities of staphylococcal enterotoxin A (SEA), Car boxymethylated SEA was unable to stimulate murine T-cell proliferation but was resistant to monkey stomach lavage fluid degradation, suggest ing that native conformation was intact. Site-directed mutagenesis of the histidine residues of SEA was subsequently performed, SEA-H44A (SE A with histidine 44 replaced with alanine), SEA-H44D, SEA-H50A, SEA-H5 0D, SEA-H114A, SEA-H114D, SEA-H187A, and SEA-H187D retained superantig en and emetic activities, whereas SEA-H225A and SEA-H225D were defecti ve in the ability to stimulate T-cell proliferation. These mutants wer e unable to compete with SEA for binding to Raji cells, suggesting tha t the defect in SEA-H225A and SEA-H225D is due to impaired major histo compatibility complex class II binding. SEA-H225D provoked an emetic r esponse in monkeys only if fed at high doses, while SEA-H225A did not provoke an emetic response at low or high doses. In comparison, SEA-H6 1A and SEA-H61D were defective in emetic activity but not in the abili ty to stimulate murine T-cell proliferation. Overall, these studies sh ow that the carboxy-terminal histidine at residue position 225 of SEA is important for both the superantigen and emetic activities of this e nterotoxin. Histidine 61 appears to be important for emetic activity b ut not for superantigen activity, consistent with the hypothesis that the two activities are separable in staphylococcal enterotoxins.