The ERAP1 active site cannot productively access the N-terminus of antigenic peptide precursors stably bound onto MHC class I

George Mavridis,Dimitris Georgiadis,Anastasia Mpakali,Jerome Zoidakis,Eleni Kaloumenou,Manousos Makridakis,Athanasios Papakyriakou,Angeliki Ziotopoulou,Antonia Vlahou,Efstratios Stratikos

doi:10.1038/s41598-021-95786-x

Abstract

Processing of N-terminally elongated antigenic peptide precursors by Endoplasmic Reticulum Aminopeptidase 1 (ERAP1) is a key step in antigen presentation and the adaptive immune response. Although ERAP1 can efficiently process long peptides in solution, it has been proposed that it can also process peptides bound onto Major Histocompatibility Complex I molecules (MHCI). In a previous study, we suggested that the occasionally observed “ontο MHCI” trimming by ERAP1 is likely due to fast peptide dissociation followed by solution trimming, rather than direct action of ERAP1 onto the MHCI complex. However, other groups have proposed that ERAP1 can trim peptides covalently bound onto MHCI, which would preclude peptide dissociation. To explore this interaction, we constructed disulfide-linked MHCI-peptide complexes using HLA-B*08 and a 12mer kinetically labile peptide, or a 16mer carrying a phosphinic transition-state analogue N-terminus with high-affinity for ERAP1. Kinetic and biochemical analyses suggested that while both peptides could access the ERAP1 active site when free in solution, they were unable to do so when tethered in the MHCI binding groove. Our results suggest that MHCI binding protects, rather than promotes, antigenic peptide precursor trimming by ERAP1 and thus solution trimming is the more likely model of antigenic peptide processing.

Highlights

Processing of N-terminally elongated antigenic peptide precursors by Endoplasmic Reticulum Aminopeptidase 1 (ERAP1) is a key step in antigen presentation and the adaptive immune response
In a recent paper from our group, we examined the kinetics of ERAP1-trimming of antigenic peptide precursors bound onto three different MHCI alleles and concluded that in all cases MHCI protected the peptides from ERAP1 trimming, suggesting that solution trimming is the main mechanism of ERAP1-mediated antigenic peptide generation[17]
We have previously demonstrated that the 12mer peptide with the sequence ARAALRSRYWAI can be refolded with HLA-B*08:01 to form a thermodynamically stable but kinetically labile complex and that ERAP1 can apparently trim this peptide while it is bound onto HLA-B*08:01, albeit slower than in solution[17]

Summary

Introduction

Processing of N-terminally elongated antigenic peptide precursors by Endoplasmic Reticulum Aminopeptidase 1 (ERAP1) is a key step in antigen presentation and the adaptive immune response. To further explore whether it is necessary for the peptide to dissociate from HLA-B*08 before being trimmed by ERAP1, as suggested p reviously[17], we constructed a disulfide-linked version of the complex using the E76C mutation in HLA-B*08 that introduces a cysteine residue in the binding groove of the M HCI18.

Results

Conclusion