The Internal Sequence of the Peptide-Substrate Determines Its N-Terminus Trimming by ERAP1

Irini Evnouchidou,Athanasios Papakyriakou,Leondios Leondiadis,Angeliki Chroni,Efstratios Stratikos,Shih-Chung Chang,Frank Momburg,Alfred L Goldberg,Hany A El-Shemy

doi:10.1371/journal.pone.0003658

Abstract

BackgroundEndoplasmic reticulum aminopeptidase 1 (ERAP1) trims N-terminally extended antigenic peptide precursors down to mature antigenic peptides for presentation by major histocompatibility complex (MHC) class I molecules. ERAP1 has unique properties for an aminopeptidase being able to trim peptides in vitro based on their length and the nature of their C-termini.Methodology/Principal FindingsIn an effort to better understand the molecular mechanism that ERAP1 uses to trim peptides, we systematically analyzed the enzyme's substrate preferences using collections of peptide substrates. We discovered strong internal sequence preferences of peptide N-terminus trimming by ERAP1. Preferences were only found for positively charged or hydrophobic residues resulting to trimming rate changes by up to 100 fold for single residue substitutions and more than 40,000 fold for multiple residue substitutions for peptides with identical N-termini. Molecular modelling of ERAP1 revealed a large internal cavity that carries a strong negative electrostatic potential and is large enough to accommodate peptides adjacent to the enzyme's active site. This model can readily account for the strong preference for positively charged side chains.Conclusions/SignificanceTo our knowledge no other aminopeptidase has been described to have such strong preferences for internal residues so distal to the N-terminus. Overall, our findings indicate that the internal sequence of the peptide can affect its trimming by ERAP1 as much as the peptide's length and C-terminus. We therefore propose that ERAP1 recognizes the full length of its peptide-substrate and not just the N- and C- termini. It is possible that ERAP1 trimming preferences influence the rate of generation and the composition of antigenic peptides in vivo.

Highlights

Antigenic peptides presented by major histocompatibility complex (MHC) class I molecules act as a status indicator of the cell’s condition and play a pivotal role in the activation of T-lymphocytes versus pathogens like viruses or in pathological conditions like cancer
We found that Endoplasmic reticulum aminopeptidase 1 (ERAP1) exhibited very strong preferences for specific amino acids in several positions of the peptide substrate, for hydrophobic and positively charged side-chains
In this study we used collections of model peptides to systematically study the effects of peptide sequence in the degradation of its Nterminus by ERAP1

Summary

Introduction

Antigenic peptides presented by MHC class I molecules act as a status indicator of the cell’s condition and play a pivotal role in the activation of T-lymphocytes versus pathogens like viruses or in pathological conditions like cancer. The proteasomegenerated peptides destined for MHC loading generally have the correct C-terminus for MHC binding but carry N-terminal extensions that need to be trimmed away [20]. This trimming is completed in the ER it may be initiated in the cytosol. The immunoproteasome generates N-terminal extended antigenic peptide precursors more efficiently than the proteasome, something that may help the peptides survive the proteolytic activity of the cytosol and enhance their chances to be transported into the ER [20]. Endoplasmic reticulum aminopeptidase 1 (ERAP1) trims N-terminally extended antigenic peptide precursors down to mature antigenic peptides for presentation by major histocompatibility complex (MHC) class I molecules. ERAP1 has unique properties for an aminopeptidase being able to trim peptides in vitro based on their length and the nature of their C-termini

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Conclusion