The polymorphism at position beta69 of the
human leukocyte antigen (
HLA)-DP molecule has been associated with susceptibility to several
immune disorders and alloreactivity. Using molecular modeling, we have predicted a detailed structure of the
HLA-DP2 molecule (carrying Glubeta69) complexed with
class II associated invariant chain derived
peptide (
CLIP) and compared it with the form carrying Lys at beta69 (HLA-DP2K69). Major changes between the two models were observed in the shape and charge distribution of pocket 4 and of the nearby pocket 6. Consequently, we analyzed in detail the
peptide-binding specificities of both
HLA-DP molecules expressed as
recombinant proteins. We first determined that the minimum
peptide-binding core of
CLIP for both
HLA-DP2 and DP2K69 is represented by nine aminoacids corresponding to the sequence 91-99 of
invariant chain (Ii). We then assessed the
peptide-binding specificities of the two pockets and determined the role of position beta69, using competition tests with the Ii-derived
peptide CLIP and its mutated forms carrying all the aminoacidic substitutions in P4 and P6. Pocket 4 of
HLA-DP2 showed high affinity for positively charged, aromatic, and polar residues, whereas aliphatic residues were disfavored. Pocket 4 of the DP2K69 variant showed a reduced aminoacid selectivity with aromatic residues most preferred. Pocket 6 of
HLA-DP2 showed high affinity for aromatic residues, which was increased in DP2K69 and extended to
arginine. Finally, we used the experimental data to determine the best molecular-modeling approach for assessing aminoacid selectivity of the two pockets. The results with best predictive value were obtained when single aminoacids were evaluated inside each single pocket, thus, reducing the influence of the overall
peptide/ major histocompatibility complex interaction. In conclusion, the HLA-DPbeta69 polymorphism plays a fundamental role in the
peptide-binding selectivity of
HLA-DP. Furthermore, as this polymorphism is the main change in the pocket 4 area of
HLA-DP, it could represent a supertype among
HLA-DP molecules significantly contributing to the selection of
epitopes presented in the context of this HLA isotype.