A systematic re-examination of processing of MHCI-bound antigenic peptide precursors by endoplasmic reticulum aminopeptidase 1

Abstract

Endoplasmic reticulum aminopeptidase 1 (ERAP1) trims antigenic peptide precursors to generate mature antigenic peptides for presentation by major histocompatibility complex class I (MHCI) molecules and regulates adaptive immune responses. ERAP1 has been proposed to trim peptide precursors both in solution and in pre-formed MHCI-peptide complexes, but which mode is more relevant to its biological function remains controversial. Here, we compared ERAP1-mediated trimming of antigenic peptide precursors in solution or when bound to three MHCI alleles, HLA-B*58, HLA-B*08 and HLA-A*02. For all MHCIpeptide combinations, peptide binding onto MHCI protected against ERAP1-mediated trimming. In only a single MHCI-peptide combination, trimming of an HLA-B*08-bound 12mer progressed at a considerable rate, albeit still slower than in solution. Results from thermodynamic, kinetic and computational analyses suggested that this 12mer is highly labile and that apparent on-MHC trimming rates are always slower than that of MHCI-peptide dissociation. Both ERAP2 and leucine aminopeptidase, an enzyme unrelated to antigen processing, could trim this labile peptide from pre-formed MHCI complexes as efficiently as ERAP1. A pseudopeptide analogue with high affinity for both HLA-B*08 and the ERAP1 active site could not promote the formation of a ternary ERAP1-MHCI-peptide complex. Similarly, no interactions between ERAP1 and purified peptide loading complex (PLC) were detected in the absence or presence of a pseudopeptide trap. We conclude that MHCI binding protects peptides from ERAP1 degradation and that trimming in solution, along with the dynamic nature of peptide binding to MHCI, are sufficient to explain ERAP1 processing of antigenic peptide precursors.

ERAP2 increases the abundance of a peptide submotif highly selective for the Birdshot Uveitis-associated HLA-A29

“…Both ERAP1 and ERAP2 have been shown to have preferences for the internal sequence of the peptide, although these preferences are broad and no specific motif has been identified 13–16 . These and other observations 17 support that ERAPs predominantly modulate the ‘free’ peptide cargo before binding to HLA, which suggests that physiologically-relevant sequence specificities for ERAP2 may be deciphered from the presented peptide repertoire.…”

“…Both ERAP1 and ERAP2 have been shown to have preferences for the internal sequence of the peptide, although these preferences are broad and no specific motif has been identified 13–16 . However, some studies have also shown that ERAPs can also trim peptide while they are bound onto MHC-I 17,18 , although this mechanism has been brought into question 19 . These and other observations 19 support that ERAPs modulate a significant fraction of the ‘free’ peptide cargo before binding to HLA, which suggests that physiologically-relevant sequence specificities for ERAP2 may be deciphered from the presented peptide repertoire.…”

ERAP2 Increases the Abundance of a Peptide Submotif Highly Selective for the Birdshot Uveitis-Associated HLA-A29

“…However, a number of studies have also shown that ERAPs can also trim peptide while they are bound onto MHC-I ( 17 – 21 ). These and other observations ( 22 ) support that ERAPs modulate both HLA-bound and a significant fraction of the ‘free’ peptide cargo before binding to HLA, which suggests that physiologically-relevant sequence specificities for ERAP2 may be deciphered from the presented peptide repertoire.…”

Generation of SARS-CoV-2 S1 Spike Glycoprotein Putative Antigenic Epitopes in Vitro by Intracellular Aminopeptidases

“… 43 Further systematic analysis of this phenomenon however, suggested that it is either very rare or misinterpreted in the case of rapid-dissociating peptides. 44 We therefore did not attempt to characterize trimming of prebound peptides in this study.…”