Large Scale Mass Spectrometric Profiling of Peptides Eluted from HLA Molecules Reveals N-Terminal-Extended Peptide Motifs

Abstract

The majority of >2000 HLA class I molecules can be clustered according to overlapping peptide binding specificities or motifs recognized by CD8 ؉ T cells. HLA class I motifs are classified based on the specificity of residues located in the P2 and the C-terminal positions of the peptide. However, it has been suggested that other positions might be relevant for peptide binding to HLA class I molecules and therefore be used for further characterization of HLA class I motifs. In this study we performed large-scale sequencing of endogenous peptides eluted from K562 cells (HLA class I null) made to express a single HLA molecule from HLA-B*3501, -B*3502, -B*3503, -B*3504, -B*3506, or -B*3508. Using sequence data from >1,000 peptides, we characterized novel peptide motifs that include dominant anchor residues extending to all positions in the peptide. The length distribution of HLA-B35-bound peptides included peptides of up to 15 residues. Remarkably, we determined that some peptides longer than 11 residues represented N-terminal-extended peptides containing an appropriate HLA-B35 peptide motif. These results provide evidence for the occurrence of endogenous N-terminal-extended peptide-HLA class I configurations. In addition, these results expand the knowledge about the identity of anchor positions in HLA class I-associated peptides that can be used for characterization of HLA class I motifs.

Expanding the detectable HLA peptide repertoire using electron-transfer/higher-energy collision dissociation (EThcD)

“…We identified a large set of nested peptides that differ in length either at the N or C terminus. The presence of N-terminal-extended length variants have been reported previously (17,32). The activity of aminopeptidases in the ER effectively edits the majority of endogenously processed peptides to a length suitable for loading onto the HLA class I molecules, but a minority of peptides escape this efficient trimming process (33).…”

Analysis of Major Histocompatibility Complex-Bound HIV Peptides Identified from Various Cell Types Reveals Common Nested Peptides and Novel T Cell Responses

“…This MHC-peptidome study and others in viral infections (9,15,16) or in B cell lines (30,(42)(43)(44)(45) identified clusters of nested peptides. They include optimal epitopes as well as N-and/or Cextended peptides potentially presentable by several HLAs.…”

Peptide-dependent Conformational Fluctuation Determines the Stability of the Human Leukocyte Antigen Class I Complex

“…Nevertheless, both peptides were bound to HLA-B*35:01 in the typical manner in which the B and F pockets prefer peptides with a proline residue at position 2 and either a tyrosine, phenylalanine, methionine, or leucine residue at the carboxyl terminus (40,41). The remaining pockets (A, C, D, and E) exhibit little preference with respect to the amino acid sequence of the bound peptide (12,13). The second proline and the carboxyl-terminal tyrosine in both VY8 and RY11 form hydrogen bonds with Tyr-99 in the B pocket and Ser-116 in the F pocket, respectively ( Fig.…”

“…The heavy chain contains the peptide binding domain, with six pockets (A-F pockets) that vary in structure among different HLA class I allomorphs. For most HLA allomorphs, two of the pockets play a dominant role in determining peptide specificity (12,13). The antigenic surface formed by the bound peptide and the peptide binding domain of the heavy chain is unique to each pHLA structure.…”