FUNCTIONAL-ACTIVITY OF STAPHYLOCOCCAL-ENTEROTOXIN-A REQUIRES INTERACTIONS WITH BOTH THE ALPHA-CHAIN AND BETA-CHAIN OF HLA-DR

The staphylococcal enterotoxins, SEA and SEE, bind one zinc atom per m olecule of protein. The presence of this metal atom enhances the bindi ng of the toxins to MHC class II molecules, presumably through an inte raction with histidine 81 of the beta chain. L cell transfectants expr essing HLA-DR1 and HLA-DR7 molecules, with mutations in either the alp ha 1 or beta 1 domains, were tested for their ability to bind SEA and present it to T cells. Cells expressing DR1 molecules with alanine at positions 77, 78, 80, 83, 84 and 85, or serine at position 79 could al l bind SEA and present it to either polyclonal or monoclonal T cells. Most point mutations within the alpha-helical portion of the DR7 beta chain had no effect on binding and presentation. However, substitution of histidine 81 with alanine, glutamate, or aspartate, abrogated SEA binding as well as T cell stimulation by the superantigen. This effect was also observed when the non-polymorphic aspartate, at position 76 was changed to alanine. Mutation of the asparagine at position 82 had an intermediate effect. Point mutations of the DR alpha chain had litt le effect on binding of SEA as determined by a flow cytometric assay. However, mutation of lysine at position 39 of the alpha chain and, to a lesser extent methionine at position 36, markedly decreased the abil ity of SEA to stimulate toxin-responsive mouse T cell hybridomas. Fina lly, the monoclonal antibody, L243 binds to the alpha chain of HLA-DR, and was able to block T cell activation by SEA without blocking SEA b inding. These data support the model whereby HLA-DR has two binding si tes for SEA. A low affinity site, present on the alpha chain, is requi red for T cell stimulation by the superantigen, but is insufficient to mediate toxin binding. High affinity binding of HLA-DR to SEA occurs solely through residues on the beta chain, including both histidine 81 and aspartate 76. (C) 1997 Elsevier Science Ltd.