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Related Experiment Video

Updated: May 20, 2026

LERLIC-MS/MS for In-depth Characterization and Quantification of Glutamine and Asparagine Deamidation in Shotgun Proteomics
08:01

LERLIC-MS/MS for In-depth Characterization and Quantification of Glutamine and Asparagine Deamidation in Shotgun Proteomics

Published on: April 9, 2017

Top-down study of β2-microglobulin deamidation.

Xiaojuan Li1, Xiang Yu, Catherine E Costello

  • 1Mass Spectrometry Resource, Department of Biochemistry, Boston University School of Medicine, Boston, Massachusetts 02118, United States.

Analytical Chemistry
|July 4, 2012
PubMed
Summary
This summary is machine-generated.

Identifying isoAspartic acid (isoAsp) in intact proteins is challenging. A new MS(3) method combining CAD-ECD successfully detected isoAsp at all sites in beta2-microglobulin, enabling quantitative analysis.

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Last Updated: May 20, 2026

LERLIC-MS/MS for In-depth Characterization and Quantification of Glutamine and Asparagine Deamidation in Shotgun Proteomics
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Sample Preparation and Relative Quantitation using Reductive Methylation of Amines for Peptidomics Studies
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Sample Preparation and Relative Quantitation using Reductive Methylation of Amines for Peptidomics Studies

Published on: November 4, 2021

Area of Science:

  • Mass spectrometry
  • Proteomics
  • Biochemistry

Background:

  • Distinguishing aspartic acid (Asp) and isoAspartic acid (isoAsp) isomers from Asn deamidation is crucial for protein analysis.
  • While Electron Capture Dissociation (ECD) excels at peptide-level isomer differentiation, intact protein analysis remains difficult.
  • Beta2-microglobulin (β(2)M) serves as a model system for studying protein deamidation.

Purpose of the Study:

  • To develop and validate a top-down mass spectrometry approach for identifying isoAsp formation in intact proteins.
  • To overcome limitations of standard ECD for isoAsp detection in large molecules.
  • To enable quantitative analysis of isoAsp formation in aged proteins.

Main Methods:

  • Top-down mass spectrometry was employed using beta2-microglobulin (β(2)M) as a model.
  • Initial top-down ECD analysis was performed on intact, deamidated β(2)M.
  • An MS(3) strategy involving Collisionally Activated Dissociation (CAD) followed by ECD was developed and applied.

Main Results:

  • Standard top-down ECD identified isoAsp at only one of three deamidation sites in β(2)M.
  • The MS(3) CAD-ECD approach successfully identified isoAsp formation at all three deamidation sites.
  • A linear correlation was observed between the abundance of the isoAsp diagnostic ion and the deamidation extent.

Conclusions:

  • The MS(3) CAD-ECD approach significantly enhances the capability of top-down mass spectrometry for isoAsp detection in intact proteins.
  • This method overcomes previous limitations in differentiating deamidation isomers at the intact protein level.
  • The findings pave the way for improved quantitative analysis of protein deamidation and aging.