The P9 pocket of HLA-DQ2 (non-Aspbeta57) has no particular preference for negatively charged anchor residues found in other type 1 diabetes-predisposing non-Aspbeta57 MHC class II molecules.

Susceptibility and resistance to type 1 diabetes are associated with MHC class II alleles that carry non-Asp and Asp at residue 57 of their b chain respectively. The effect of Asp or non-Aspb57 may relate to a differential ability of distinct class II molecules to bind specific immuno-pathogenic peptides. Recent studies in man and mouse have revealed that some type 1 diabetes-predisposing non-Aspb57 class II molecules (i.e. DQ8, DR4Dw15 and I-A g7 ) preferentially bind peptides with a negatively charged anchor residue at P9. It has been suggested that this is a common feature of type 1 diabetes-predisposing class II molecules. The molecular explanation for such a phenomenon could be that class II b chains with Aspb57 form a salt bridge between Aspb57 and a conserved Arg of the a chain, whereas in non-Aspb57 molecules the Arg is unopposed and free to interact with negatively charged P9 peptide anchor residues. We have investigated the specificity of the P9 pocket of the type 1 diabetes-associated DQ2 molecule and in particular examined for charge effects at this anchor position. Different approaches were undertaken. We analyzed binding of a high-affinity binding ligand and P9-substituted variants of this peptide, and we analyzed the binding of a set of synthetic random peptide libraries. The binding analyses were performed with wild-type DQ2 and a mutated DQ2 with Ala at b57 substituted with Asp. Our results indicate that the wild-type DQ2 (non-Aspb57) prefers large hydrophobic residues at P9 and that there is no particular preference for binding peptides with negatively charged residues at this position. The specificity of the P9 pocket in the mutated DQ molecule is altered, indicating that the b57 residue contributes to determining the specificity of the P9 pocket. Our data do not lend support to the hypothesis that all non-Asp b57 class II molecules predispose to development of disease by binding peptides with negatively charged P9 anchor residues.