EFFECTS OF MHC-II CONFORMATION AND PH ON THE RECOGNITION OF PEPTIDE BY T-CELLS

In this study we analysed the binding of the peptide HEL46-61 to purif ied A(k) molecules which have been altered by site-directed mutagenesi s at polymorphic positions to include amino acids from the A(d) alpha- chain. We find that changes in the floor of the peptide binding groove , at positions 11, 14 and 28, abolish T cell recognition without chang ing peptide binding affinity. We further show that amino acid changes at these positions in the A(d) molecule result in a conformationally a ltered molecule as evidenced by loss of binding of the A(alpha)(d) spe cific monoclonal antibody K24. Thus the T cell receptor is highly sens itive to subtle changes in MHC II structure induced at sites that are unlikely to be involved in direct T cell contact. This has important i mplications with respect to allorecognition. The binding studies repor ted here were performed both at pH7 7, to reflect binding of peptides at the cell surface, and at pH 5.5, to mimic binding in an intracellul ar acidic compartment. Binding to wild-type A(k) was increased 2-3-fol d at pH 5.5, whereas binding to some MHC II mutants was increased by g reater than 20-fold at pH 5.5 relative to pH 7. These results show tha t the apparent peptide binding specificity for the mutants differs at pH 7 and 5.5, and suggest that caution should be used in defining the MHC-restriction of peptide epitopes at neutral pH.