OR16: CHARACTERIZATION THE MAJOR AND MINOR LIGAND COMPARTMENT OF HLA-E

Aim HLA-E is a non-classical class I HLA that binds and presents class I MHC leader sequences which serve as a ligand for NK receptors such as CD94/NKG2. Apart from this function HLA-E has been reported to bind HLA-A2 like peptides in the absence of TAP. In this study we aim to identify the endogenous ligands presented by HLA-E in the presence of class I with the full complement of chaperones. Methods Here we collect purified soluble HLA-E*01:03 from the U373 cell line. Using acid elution we isolate the peptide ligand pool and analyze them using 2-dimentional LCMS. MS spectra are collected using information-dependant acquisition on a Triple-TOF AB Sciex 5600. Fragment spectra are analyzed with PEAKS and Mascot using the decoy database method at a 1%FDR. Results As initially reported for HLA-E, leader peptides derived from classical HLA, are predominant peptide ligands. We also observed multiple length variants of the leader peptides as well post-translationally modified variants. In addition, greater than 1500 host derived non-leader ligands from 389 different proteins were presented by HLA-E, demonstrating a greater scope of peptide sampling than initially suspected for HLA-E. Within these minor ligands, we observed a bi-modal distribution in the length of the peptides. Consistent with previous reports, ligands 8–11 amino acids in length (N = 597) show a motif similar to HLA-A2. However ligands with >11 amino acids (N = 1035) demonstrate a different motif where the P2 is L,V,G, PΩ is D,K,R and PΩ-1 is L. Conclusion In summary, we show that even in the presence of HLA leader peptides, HLA-E binds a large diversity of peptide ligands with two distinct length preferences and motifs.