Effect of the Asn side chain on the dissociation of deprotonated peptides elucidated by IRMPD spectroscopy

Abstract

Dedication: This paper is dedicated to the memory of our dear friend Detlef Schröder. Table of contents graphic Highlights • Non-oxazolone b-type fragments identified for deprotonated peptides • b-type anions from Asn-containing peptides have succinimide structure • b 2 and b 3 anions investigated• IR spectroscopy distinguishes between imide and isoimide structures Abstract Infrared ion spectroscopy using the free electron laser FELIX was applied to identify the structure of b-type peptide fragments generated by collision and IR multiple-photon induced dissociation from singly deprotonated peptides containing an asparagine residue, in particular AlaAsnAla (ANA) and AlaAlaAsnAla (AANA). IR spectra were recorded over the 800-1800 cm -1 spectral range by multiple-photon dissociation (IRMPD) spectroscopy and have been compared with density functional theory (DFT) calculated spectra at the B3LYP/6-31++G(d,p) level for different isomeric ion structures for structural characterization. Results unambiguously show that the b 2 and b 3 fragment anions do not possess the common oxazolone or diketopiperazine structure, but involve cyclization of the asparagine side chain. Nucleophilic attack from the side chain amide nitrogen on the peptide backbone carbonyl carbon leads to the formation of cyclic succinimide structures. Deprotonation is shown to occur on the succinimide nitrogen, which delocalizes the negative charge over two adjacent carbonyl groups thus enhancing the gasphase stability.

Dehydration reactions of protonated dipeptides containing asparagine or glutamine investigated by infrared ion spectroscopy

“…In its most common form, dissociation is accomplished through collisional activation. Collision-induced dissociation (CID) primarily leads to backbone dissociation at the amide bonds, generating sequence ions referred to as b-type (N-terminal) or y-type (C-terminal) [1][2][3][4][5][6][7][8]. The molecular structures of the resulting b-type ions have been under particularly vigorous debate and it has now been established that b-type ions most often possess an oxazolone structure; for b2-fragment ions, a diketopiperazine structure is in some cases formed alternatively [9].…”

Ion Mobility Spectrometry-Hydrogen Deuterium Exchange Mass Spectrometry of Anions: Part 2. Assessing Charge Site Location and Isotope Scrambling

“…The extent of HD scrambling depends not only upon the effective temperature of the ions but also the differences in energy levels of different charge configurations [24]. The same proton mobilization process is required for peptide anion fragmentation [25]. One difference between positive-ion and negative-ion analysis is the source of protons.…”

Deamidation reactions of protonated asparagine and glutamine investigated by ion spectroscopy

“…A variety of studies have addressed the deamidation reactions of protonated Gln-and Asn-containing proteins and peptides in a mass spectrometer, where the reactions are initiated by collisional activation, occur in the gas phase, and can be analyzed in terms of the mass-to-charge ratio of the resulting fragments. [1,3,6,[12][13][14][15][16][17][18][19]31,32] In order to gain detailed mechanistic insight into these reactions, the deamidation of protonated Asn and Gln has been studied using a variety of methods including energy-dependent collision-induced dissociation, deuterium exchange, and threshold collision-induced dissociation (TCID) combined with theoretical investigations at various levels of theory.…”