Effect of charge derivatization in the determination of phosphorylation sites in peptides by electrospray ionization collision-activated dissociation tandem mass spectrometry

Sadagopan, Nalini; Malone, Michael J.; Watson, J. Throck

doi:10.1002/(sici)1096-9888(199912)34:12<1279::aid-jms899>3.0.co;2-9

Cited by 6 publications

(2 citation statements)

References 26 publications

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“…Derivatisation strategies have been developed to circumvent some properties of the phosphate moiety, such as the loss of phosphoric acid under CID conditions, and bad ionisation behaviour [171,172,173,174]. β-elimination and Michael addition (Fig.…”

Section: Derivatisation Of Phosphopeptidesmentioning

confidence: 99%

The impact of chromatography and mass spectrometry on the analysis of protein phosphorylation sites

Zeller

König

2004

Analytical and Bioanalytical Chemistry

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Protein phosphorylation analysis is an enormous challenge. This review summarises the currently used techniques, which are based on radiolabelling and mass spectrometry as well as electrophoretic and chromatographic separation. Many methods exist, but there is still no single procedure applicable to all phosphoproteins. MS is able to deliver information about the location of phosphorylation sites, but phosphospecific properties with respect to ionisation present obstacles. Therefore, multidimensional approaches involving several analytical methods are often necessary to conquer phosphorylation site identification.

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Section: Derivatisation Of Phosphopeptidesmentioning

confidence: 99%

The impact of chromatography and mass spectrometry on the analysis of protein phosphorylation sites

Zeller

König

2004

Analytical and Bioanalytical Chemistry

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“…In an alternative approach, phosphate [19] or glycan moieties [20] have been localized by CAD-MS/MS or postsource decay analysis of peptides derivatized at their N-termini with a phosphonium group. It has been shown that when a singly charged cation was selected as an ion precursor, loss of the conjugated moiety could be almost completely suppressed.…”

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confidence: 99%

The combination of electron capture dissociation and fixed charge derivatization increases sequence coverage for O-glycosylated and O-phosphorylated peptides

Chamot‐Rooke

Rest

Dalleu

et al. 2007

J. Am. Soc. Mass Spectrom.

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Electron capture dissociation (ECD) has become an alternative method to collision-activated dissociation (CAD) to avoid gas-phase cleavage of post-translational modifications carried by side chains from the peptide backbone. Nonetheless, as illustrated herein by the study of O-glycosylated and O-phosphorylated peptides, the extent of ECD fragmentations may be insufficient to cover the entire peptide sequence and to localize accurately these modifications. The present work demonstrates that the derivatization of peptides at their N-terminus by a phosphonium group improves dramatically and systematically the sequence coverage deduced from the ECD spectrum for both O-glycosylated and O-phosphorylated peptides compared with their native counterparts. The exclusive presence of N-terminal fragments (c-type ions) in the ECD spectra of doubly charged molecular cations simplifies peptide sequence interpretation. Thus, the combination of ECD and fixed charge derivatization appears as an efficient analytical tool for the extensive sequencing of peptides bearing labile groups. (J Am Soc Mass Spectrom 2007, 18, 1405-1413) © 2007 American Society for Mass Spectrometry T he field of proteomics usually aims at differential characterization of the complete pattern of protein expression, for instance between cells in a normal state and those in a pathological state. Protein identification is now routinely achieved on a large scale (tens or hundreds of proteins per day) by combining enzymatic digestion, mass spectrometric analysis of the resulting peptides, and subsequent database search. If few peptides or short peptide sequences are sufficient for protein identifications into databases, a complete determination of the primary structure is nevertheless useful to detect modifications that can be involved in the regulation of protein function. Indeed, the catalytic property, the conformation, the turnover, or the subcellular localization of proteins are largely under the control of various peptide sequence processings named posttranslational modifications. These modifications (PTM) involve the proteolytic maturation of the proteins and/or the transfer of chemical groups (phosphorylation, glycosylation, acetylation, sumoylation) on specific amino-acids (serine, threonine, or tyrosine for O-phosphorylations, serine or threonine for Oglycosylations, asparagine for N-glycosylations). The most obvious strategy for mapping the modifications along the peptide chain involves a comparison between the sequence obtained by mass spectrometric data and the one deduced from the open reading frame coded by the gene sequence. However, a drawback arises from the propensity of groups conjugated to the amino acid side chains to dissociate readily upon collisional activation. For instance, collisionally-activated dissociation (CAD) MS/MS spectra of protonated precursor ions of phosphorylated peptides exhibit intense ions originating from the elimination of metaphosphoric (HPO 3 , loss of 80 Da) or orthophosphoric acid (H 3 PO 4 , loss of 98 Da) [1]. I...

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Mass Spectrometry: Structure Determination of Proteins and Peptides☆

Zoidakis¹,

Chatziharalambous²

2016

Reference Module in Chemistry, Molecular Sciences and Chemical Engineering

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Effect of charge derivatization in the determination of phosphorylation sites in peptides by electrospray ionization collision-activated dissociation tandem mass spectrometry

Cited by 6 publications

References 26 publications

The impact of chromatography and mass spectrometry on the analysis of protein phosphorylation sites

The impact of chromatography and mass spectrometry on the analysis of protein phosphorylation sites

The combination of electron capture dissociation and fixed charge derivatization increases sequence coverage for O-glycosylated and O-phosphorylated peptides

Mass Spectrometry: Structure Determination of Proteins and Peptides☆

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