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Related Experiment Video

Updated: Mar 8, 2026

Quantification of Site-specific Protein Lysine Acetylation and Succinylation Stoichiometry Using Data-independent Acquisition Mass Spectrometry
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Quantification of Site-specific Protein Lysine Acetylation and Succinylation Stoichiometry Using Data-independent Acquisition Mass Spectrometry

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Site-Specific Quantification of Lysine Acetylation Using Isotopic Labeling.

M Miyagi1

  • 1Center for Proteomics and Bioinformatics, Case Western Reserve University, Cleveland, OH, United States.

Methods in Enzymology
|February 1, 2017
PubMed
Summary

This study introduces a novel method to quantify site-specific lysine acetylation stoichiometry. The technique uses chemical labeling and mass spectrometry to reveal the extent of protein acetylation at individual sites.

Keywords:
AcetylationIsotope labelingLC-MS/MSMass spectrometryProteomicsStoichiometry

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Area of Science:

  • Biochemistry
  • Proteomics
  • Molecular Biology

Background:

  • Lysine acetylation is a crucial, reversible post-translational modification regulating diverse cellular functions.
  • Understanding site-specific acetylation stoichiometry is vital for elucidating its functional roles.
  • Current methods lack the precision to quantify acetylation levels at individual lysine sites.

Purpose of the Study:

  • To develop and validate a method for determining site-specific lysine acetylation stoichiometry.
  • To enable quantitative analysis of acetylation dynamics in various physiological contexts.
  • To facilitate a deeper understanding of the functional impact of site-specific acetylation.

Main Methods:

  • A chemical labeling strategy using isotopically labeled acetyl groups (¹³C₂-acetyl) to tag unacetylated lysines.
  • Subsequent mass spectrometry analysis to compare endogenous (¹²C₂) and labeled (¹³C₂) acetyl groups.
  • Application to pure proteins or simple protein mixtures for accurate stoichiometry determination.

Main Results:

  • The described method accurately quantifies the stoichiometry of lysine acetylation at specific sites.
  • It allows differentiation between endogenous and chemically introduced acetyl modifications.
  • The approach is effective for analyzing acetylation levels in purified proteins.

Conclusions:

  • This method provides a robust tool for quantitative proteomics, specifically for lysine acetylation.
  • It opens new avenues for investigating the functional significance of site-specific acetylation.
  • The technique is valuable for studying dynamic changes in protein acetylation under different conditions.