The Impact of Phosphorylation on Electron Capture Dissociation of Proteins: A Top-Down Perspective

The Impact of Phosphorylation on Electron Capture Dissociation of Proteins: A Top-Down Perspective Electron capture dissociation (ECD) is well suited for the characterization of phosphoproteins, with which labile phosphate groups are generally preserved during the fragmentation process. However, the impact of phosphorylation on ECD fragmentation of intact proteins remains unclear. Here, we have performed a systematic investigation of the phosphorylation effect on ECD of intact proteins by comparing the ECD cleavages of mono-phosphorylated α-casein, multi-phosphorylated β-casein, and immunoaffinity-purified phosphorylated cardiac troponin I with those of their unphosphorylated counterparts, respectively. In contrast to phosphopeptides, phosphorylation has significantly reduced deleterious effects on the fragmentation of intact proteins during ECD. On a global scale, the fragmentation patterns are highly comparable between unphosphorylated and phosphorylated precursors under the same ECD conditions, despite a slight decrease in the number of fragment ions observed for the phosphorylated forms. On a local scale, single phosphorylation of intact proteins imposes minimal effects on fragmentation near the phosphorylation sites, but multiple phosphorylations in close proximity result in a significant reduction of ECD bond cleavages. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Journal of The American Society for Mass Spectrometry Springer Journals

The Impact of Phosphorylation on Electron Capture Dissociation of Proteins: A Top-Down Perspective

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Publisher
Springer US
Copyright
Copyright © 2017 by American Society for Mass Spectrometry
Subject
Chemistry; Analytical Chemistry; Biotechnology; Organic Chemistry; Proteomics; Bioinformatics
ISSN
1044-0305
eISSN
1879-1123
D.O.I.
10.1007/s13361-017-1710-3
Publisher site
See Article on Publisher Site

Abstract

Electron capture dissociation (ECD) is well suited for the characterization of phosphoproteins, with which labile phosphate groups are generally preserved during the fragmentation process. However, the impact of phosphorylation on ECD fragmentation of intact proteins remains unclear. Here, we have performed a systematic investigation of the phosphorylation effect on ECD of intact proteins by comparing the ECD cleavages of mono-phosphorylated α-casein, multi-phosphorylated β-casein, and immunoaffinity-purified phosphorylated cardiac troponin I with those of their unphosphorylated counterparts, respectively. In contrast to phosphopeptides, phosphorylation has significantly reduced deleterious effects on the fragmentation of intact proteins during ECD. On a global scale, the fragmentation patterns are highly comparable between unphosphorylated and phosphorylated precursors under the same ECD conditions, despite a slight decrease in the number of fragment ions observed for the phosphorylated forms. On a local scale, single phosphorylation of intact proteins imposes minimal effects on fragmentation near the phosphorylation sites, but multiple phosphorylations in close proximity result in a significant reduction of ECD bond cleavages.

Journal

Journal of The American Society for Mass SpectrometrySpringer Journals

Published: Jul 6, 2017

References

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