DISSOCIATION OF THE PEPTIDE-MHC CLASS-I COMPLEX LIMITS THE BINDING RATE OF EXOGENOUS PEPTIDE

Soluble, single-chain molecules for two MHC class I alleles, H-2K(d) a nd H-2K(b), were used to analyze the kinetics of antigenic peptide bin ding to MHC. After MHC preloading with radiolabeled or fluorescent pep tides, the observed rate of MHC-peptide complex dissociation increased after addition of an excess of unlabeled competitor peptide. Although exogenous peptides conforming to the allele-specific motif were requi red for the enhanced complex dissociation to occur, the dissociation r ate of the complex was independent of exogenous peptide concentration. Similarly, the association rate of exogenous peptides was independent of concentration, reflecting the presence of low affinity peptides in the binding sites of the recombinant MHC proteins; the sequences of t hese endogenous peptides conform to the consensus motif for the MHC al lele studied. Finally, the association rate of exogenous peptide decre ased when MHC molecules were preloaded with high affinity peptides, an d the binding of labeled high affinity peptide to isolated recombinant MHC was faster than the subsequent dissociation observed in the prese nce of competitor peptide. Taken together, these results imply that th e rate of exogenous peptide binding is limited by the dissociation rat e of the previously bound peptides.