A three-step kinetic mechanism for peptide binding to MHC class II proteins

Peptide binding reactions of class II MHC proteins exhibit unusual kinetics , with extremely slow apparent rate constants for the overall association ( <100 M-1 s(-1)) and dissociation (<10(-5) s(-1)) processes. Various linear and branched pathways have been proposed to account for these data. Using f luorescence resonance energy transfer between tryptophan residues in the MH C peptide binding site and aminocoumarin-labeled peptides, we measured real -time kinetics of peptide binding to empty class II MHC proteins. Our exper iments identified an obligate intermediate in the binding reaction. The obs erved kinetics were consistent with a binding mechanism that involves an in itial bimolecular binding step followed by a slow unimolecular conformation al change. The same mechanism is observed for different peptide antigens. I n addition, we noted a reversible inactivation of the empty MHC protein tha t competes with productive binding. The implications of this kinetic mechan ism for intracellular antigen presentation pathways are discussed.