Dipeptides enhance folding and peptide binding of “empty” MHC class I molecules (P5002)

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Abstract MHC (Major Histocompatibility Complex) class I molecules play a central role in the mammalian antiviral immune response by presenting the viral proteome at the cell surface for recognition by cytotoxic T lymphocytes. The folding of recombinant MHC class I molecules from denatured bacterial inclusion bodies and their assembly with the light chain beta-2 microglobulin largely depend on specific high-affinity peptides of usually eight or nine amino acids. To find the minimum peptide requirement for folding and stabilization of class I molecules, we have investigated the influence of dipeptides on the folding and peptide binding of recombinant HLA-A*02:01 (A2) and H-2Kb (Kb). In the presence of the dipeptide Gly-Leu (GL), both A2 and Kb folded well, even without a high-affinity peptide present, as measured by a tryptophan fluorescence assay. When folded in the presence of GL, they also bound much faster to their respective specific high-affinity peptides as when folded without GL. Kinetic analysis by fluorescence anisotropy revealed that the presence of GL enhances the on-rate of peptide binding to empty A2 and Kb by six to eight fold. Molecular dynamics simulation indicates that the dipeptide GL interacts with the C terminus of the peptide binding groove. In conclusion, our data demonstrate that the dipeptide GL improves the folding of A2 and Kb and induces a peptide-receptive conformation for enhanced peptide binding.