Mass spectrometric profiling of HLA-B44 peptidomes provides evidence for tapasin-mediated tryptophan editing.

Activation of CD8 + T cells against pathogens and cancers involves the recognition of antigenic peptides bound to human leukocyte antigen (HLA) class-I proteins. Peptide binding to HLA class I proteins is coordinated by a multi-protein complex called the peptide loading complex (PLC). Tapasin, a key PLC component, facilitates the binding and optimization of HLA class I peptides. However, different HLA class I allotypes have variable requirements for tapasin for their assembly and surface expression. HLA-B*44:02 and HLA-B*44:05, which differ only at residue 116 of their heavy chain sequences, fall at opposite ends of the tapasin-dependency spectrum. HLA-B*44:02 (D116) is highly tapasin-dependent, whereas HLA-B*44:05 (Y116) is highly tapasinindependent. Mass spectrometric comparisons of HLA-B*4405 and HLA-B*44:02 peptidomes were undertaken to better understand the influences of tapasin upon HLA-B44 peptidome compositions. Analyses of the HLA-B*44:05 peptidomes in the presence and absence of tapasin reveal that peptides with the C-terminal tryptophan residues and those with higher predicted binding affinities are selected in the presence of tapasin. Additionally, when tapasin is present, C-terminal tryptophans are also more highly represented among peptides unique to B*44:02 and those shared between B*44:02 and B*44:05, compared with peptides unique to B*44:05. Overall, our findings demonstrate that tapasin influences the C-terminal composition of HLA class I-bound peptides and favors the binding of higher affinity peptides. For the HLA-B44 family, the presence of tapasin or high tapasin-dependence of an allotype results in better binding of peptides with C-terminal tryptophans, consistent with a role for tapasin in stabilizing an open conformation to accommodate bulky C-terminal residues.