MUTATIONS AFFECTING THE SUPERANTIGEN ACTIVITY OF STAPHYLOCOCCAL-ENTEROTOXIN-B

As a superantigen, staphylococcal enterotoxin B (SEB) possesses the ab ility to bind to major histocompatibility complex class II molecules a nd be recognized by T cells bearing certain T-cell receptor (TCR) V be ta alleles. Other investigators have utilized site-specific mutagenesi s to generate amino acid substitutions to identify residues that may b e involved in the interaction with the TCR beta-chain. In an attempt f urther to define the face of the SEB molecule involved in the interact ion with the B-chain, we have employed a polymerase chain reaction (PC R)-based, site specific mutagenesis method to generate amino acid subs titutions with altered superantigen activity. Our results show that va line at position 169 appears ro be involved in the function of this su perantigen, since each of several substitutions at this position exhib it a significantly reduced ability to induce T-cell proliferation. Ana lysis of the responding T cells to the residue 169 substitution shows that the mutant toxins maintain TCR V beta selectivity. At the same ti me, mutation of the proximal histidine at position 166 does not alter the superantigen activity of SEB. Radiolabelled binding analysis of th ese H166 and V169 mutants shows that class II-binding activity is not significantly altered. When viewed in the context of other results rep orted in the literature, combined with the crystal structure of the to xin, our results suggest that the interaction with the TCR probably in volves SEB residues which ring a cavity along one side of the toxin mo lecule.