The partial dissociation of MHC class I bound peptides exposes their N terminus to trimming by endoplasmic reticulum aminopeptidase 1: ERAP1 trimming of MHC I-bound peptides

Abstract

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, while their C terminus is anchored at the F-pocket.

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Identifiers

ORCID for Athanasios Papakyriakou: orcid.org/0000-0003-3931-6232

ORCID for Halina Mikolajek: orcid.org/0000-0003-0776-9974

ORCID for Timothy Elliott: orcid.org/0000-0003-1097-0222

ORCID for Jorn M. Werner: orcid.org/0000-0002-4712-1833

ORCID for Edward James: orcid.org/0000-0001-8638-7928

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