Deamidation Reactions of Asparagine- and Glutamine-Containing Dipeptides Investigated by Ion Spectroscopy

Lisanne J M Kempkes,Jos Oomens,Josipa Grzetic,Jonathan Martens,Giel Berden

doi:10.1007/s13361-016-1462-5

Abstract

Deamidation is a major fragmentation channel upon activation by collision induced dissociation (CID) for protonated peptides containing glutamine (Gln) and asparagine (Asn) residues. Here, we investigate these NH3-loss reactions for four Asn- and Gln-containing protonated peptides in terms of the resulting product ion structures using infrared ion spectroscopy with the free electron laser FELIX. The influence of the side chain length (Asn versus Gln) and of the amino acid sequence on the deamidation reaction has been examined. Molecular structures for the product ions are determined by comparison of experimental IR spectra with spectra predicted by density functional theory (DFT). The reaction mechanisms identified for the four dipeptides AlaAsn, AsnAla, AlaGln, and GlnAla are not the same. For all four dipeptides, primary deamidation takes place from the amide side chain (and not from the N-terminus) and, in most cases, resembles the mechanisms previously identified for the protonated amino acids asparagine and glutamine. Secondary fragmentation reactions of the deamidation products have also been characterized and provide further insight in – and confirmation of – the identified mechanisms. Overall, this study provides a comprehensive molecular structure map of the deamidation chemistry of this series of dipeptides. Graphical ᅟ Electronic supplementary materialThe online version of this article (doi:10.1007/s13361-016-1462-5) contains supplementary material, which is available to authorized users.

Highlights

U nder natural conditions, deamidation of glutamine (Gln) and asparagine (Asn) residues in proteins is a primary route for post-translational modifications, playing a role in several diseases, and is believed to relate to aging effects acting as a molecular clock [1,2,3,4,5]
To better understand the mechanistics involved in deamidation of protonated peptides containing Gln and Asn, we employ here infrared ion spectroscopy to resolve the structures of the product ions resulting from NH3-loss from the protonated dipeptides of AlaAsn (AN), L
For the parallel reaction depicted in green in Scheme 3, the first step is a nucleophilic attack of the amide bond nitrogen on the asparagine side chain, leading to the succinimide structure 5, which was observed in the deamidation reaction of asparaginecontaining peptides in solution [3,4,5]

Summary

Introduction

U nder natural conditions, deamidation of glutamine (Gln) and asparagine (Asn) residues in proteins is a primary route for post-translational modifications, playing a role in several diseases, and is believed to relate to aging effects acting as a molecular clock [1,2,3,4,5]. The oxygen from the amide linkage can attack the asparagine side chain, leading to a 6-membered ring structure, 2, which has been proposed on the basis of a computational study for the deamidation of protonated ValAsn [35].

Results

Conclusion