Comparison of CID, ETD and metastable atom‐activated dissociation (MAD) of doubly and triply charged phosphorylated tau peptides

Abstract

The fragmentation behavior of the 2+ and 3+ charge states of eleven different phosphorylated tau peptides was studied using collision-induced dissociation (CID), electron transfer dissociation (ETD) and metastable atom-activated dissociation (MAD). The synthetic peptides studied contain up to two known phosphorylation sites on serine or threonine residues, at least two basic residues, and between four and eight potential sites of phosphorylation. CID produced mainly b-/y-type ions with abundant neutral losses of the phosphorylation modification. ETD produced c-/z-type ions in highest abundance but also showed numerous y-type ions at a frequency about 50% that of the z-type ions. The major peaks observed in the ETD spectra correspond to the charge-reduced product ions and small neutral losses from the charge-reduced peaks. ETD of the 2+ charge state of each peptide generally produced fewer backbone cleavages than the 3+ charge state, consistent with previous reports. Regardless of charge state, MAD achieved more extensive backbone cleavage than CID or ETD, while retaining the modification(s) in most cases. In all but one case, unambiguous modification site determination was achieved with MAD. MAD produced 15-20% better sequence coverage than CID and ETD for both the 2+ and 3+ charge states and very different fragmentation products indicating that the mechanism of fragmentation in MAD is unique and complementary to CID and ETD.

Ion Activation Methods for Peptides and Proteins

“…[145][146][147][148][149][150][151] For metastable atom-activated dissociation (MAD), a keV beam of helium metastable atoms interacts with protonated peptides via a combination of Penning ionization and charge reduction processes that result in fragmentation of the peptide backbone, leading to formation of a/x, b/y and c/z ions. [147][148][149] Charge transfer dissociation (CTD) uses keV helium cations to cause ionization of protonated peptides, and the exothermicity of the resulting electron abstraction process results in fragmentation of the peptides. 150 The products are ones that arise from cleavage of the C-C α backbone bond of peptides, leading to formation of a-type ion.…”

Charge Transfer Dissociation (CTD) Mass Spectrometry of Peptide Cations Using Kiloelectronvolt Helium Cations

“…Oftentimes, CID does not provide complete fragmentation of the peptide backbone and results in significant side-chain losses, including the loss of post-translational modifications, and thereby complicates the interpretation of tandem mass spectra [4,5]. These limitations have fueled a significant investment in alternative fragmentation techniques, including electron transfer dissociation (ETD) with cationic [6][7][8][9] or anionic [6,10] precursor ions, electron capture dissociation (ECD) with cationic [9,11], or anionic precursor ions [12], photodissociation [13][14][15][16][17][18][19][20], metastable atom-activated dissociation (MAD) [21][22][23][24][25][26][27][28], electron ionization dissociation (EID) [29], and electron detachment dissociation (EDD) [30]. Each technique has its merits and limitations.…”

Radical-induced fragmentation of phospholipid cations using metastable atom-activated dissociation mass spectrometry (MAD-MS)

“…An alternate activation method known as metastable atom-activated dissociation (MAD) [37–44] has been used to fragment peptides and small proteins via radical cation chemistry, and we believe this is the first report applying MAD to the fragmentation of gas-phase lipids. In contrast to CID, which almost exclusively involves even electron rearrangements, MAD causes fragmentation through radical-induced rearrangements that can induce fragmentation pathways unavailable through even electron mechanisms.…”