The partial dissociation of MHC class I–bound peptides exposes their N terminus to trimming by endoplasmic reticulum aminopeptidase 1

• Published in: The Journal of biological chemistry 2018-05, Vol.293 (20), p.7538-7548
• Main Authors: Papakyriakou, Athanasios, Reeves, Emma, Beton, Mary, Mikolajek, Halina, Douglas, Leon, Cooper, Grace, Elliott, Tim, Werner, Jörn M., James, Edward
• Format: Article
• Language: English
• Subjects: Aminopeptidases - chemistry; Aminopeptidases - metabolism; Antigen Presentation; antigen processing; Crystallography, X-Ray; Epitopes - chemistry; Epitopes - metabolism; ERAP1; free energy calculations; H2-Kb; HeLa Cells; Histocompatibility Antigens Class I - chemistry; Histocompatibility Antigens Class I - metabolism; Humans; Immunology; major histocompatibility complex (MHC); Minor Histocompatibility Antigens - chemistry; Minor Histocompatibility Antigens - metabolism; Models, Molecular; molecular dynamics; peptide trimming; Protein Conformation; Protein Domains; single chain trimer; X-ray crystallography
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.RA117.000313

Endoplasmic reticulum aminopeptidase 1 (ERAP1) and ERAP2 process N-terminally extended antigenic precursors for optimal loading onto major histocompatibility complex class I (MHC I) molecules. We and others have demonstrated that ERAP1 processes peptides bound to MHC I, but the underlying mechanism is unknown. To this end, we utilized single-chain trimers (SCT) of the ovalbumin-derived epitope SIINFEKL (SL8) tethered to the H2-Kb MHC I determinant from mouse and introduced three substitutions, E63A, K66A, and W167A, at the A-pocket of the peptide-binding groove in the MHC I heavy chain, which interact with the N termini of peptides. These variants significantly decreased SL8-presenting SCT at the cell surface in the presence of ERAP1, but did not affect overall SCT expression, indicating that ERAP1 trims the SL8 N terminus. Comparison of the X-ray crystal structures of WT and three variant SCTs revealed only minor perturbations of the peptide-binding domain in the variants. However, molecular dynamics simulations suggested that SL8 can dissociate partially within a sub-microsecond timescale, exposing its N terminus to the solvent. We also found that the C terminus of MHC I–bound SL8 remains deeply buried in the F-pocket of MHC I. Furthermore, free-energy calculations revealed that the three SCT variants exhibit lower free-energy barriers of N terminus dissociation than the WT Kb. Taken together, our results are consistent with a previously observed model in which the partial dissociation of bound peptides from MHC I exposes their N terminus to trimming by ERAP1, whereas their C terminus is anchored at the F-pocket.