A Molecular Basis for the Presentation of Phosphorylated Peptides by HLA-B Antigens.

As aberrant protein phosphorylation is a hallmark of tumor cells, the display of tumor-specific phosphopeptides by Human Leukocyte Antigen (HLA) class I molecules can be exploited in the treatment of cancer by T-cell-based immunotherapy. Yet, the characterization and prediction of HLA-I phospholigands is challenging as the molecular determinants of the presentation of such post-translationally modified peptides are not fully understood. Here, we employed a peptidomic workflow to identify 256 unique phosphorylated ligands associated with HLA-B*40, -B*27, -B*39, or -B*07. Remarkably, these phosphopeptides showed similar molecular features. Besides the specific anchor motifs imposed by the binding groove of each allotype, the predominance of phosphorylation at peptide position 4 (P4) became strikingly evident, as was the enrichment of basic residues at P1. To determine the structural basis of this observation, we carried out a series of peptide binding assays and solved the crystal structures of HLA-B*40 in complex with a phosphorylated ligand or its nonphosphorylated counterpart. Overall, our data provide a clear explanation to the common motif found in the phosphopeptidomes associated to different HLA-B molecules. The high prevalence of phosphorylation at P4 is dictated by the presence of the conserved residue Arg62 in the heavy chain, a structural feature shared by most HLA-B alleles. In contrast, the preference for basic residues at P1 is allotype-dependent and might be linked to the structure of the A pocket. This molecular understanding of the presentation of phosphopeptides by HLA-B molecules provides a base for the improved prediction and identification of phosphorylated neo-antigens, as potentially used for cancer immunotherapy.

Identification of the novel HLA allele, HLA-B*40:159, in a Taiwanese hematopoietic stem cell donor and the probable HLA haplotype in an association with B*40:159.

Using a sequence-based typing method, we found a new HLA-B*40 variant, B*40:159, in a Taiwanese hematopoietic stem cell donor. The sequence of B*40:159 is identical to the sequence of B*40:06:01:01 in exons 2 and 3 except the nucleotides at positions 412 (A → G) and 429 (A → C). The sequence variation caused two amino acid exchanges at residue 114 (N → D) and residue 116 (Y → S). The probable HLA-A, HLA-B, HLA-C and HLA-DRB1 haplotype in association with B*40:159 may be deduced as HLA-A*02:06-B*40:159-C*08:01-DRB1*08:03. The generation of B*40:159 is thought as the result of a sequence recombination event where B*46:01:01:01, B*15:01:01 or B*15:02:01 donated a minimum length of the DNA sequence from residue 412 to residue 419 to the recipient sequence of B*40:06:01:01.