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Updated: Jun 28, 2026
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Detection of Protein Ubiquitination Sites by Peptide Enrichment and Mass Spectrometry
Published on: March 23, 2020
Sulfur valence normalization to expand quantitative peptide selection in protein metrology
Wenhui Fang1,2, Manman Zhu2, Wenxin Qiu2
1Key Laboratory of Microbiological Metrology, Measurement & Bio-Product Quality Security, State Administration for Market Regulation, China Jiliang University, Hangzhou, 310018, China.
Analytical and Bioanalytical Chemistry
|June 26, 2026
Summary
This study presents a new method to accurately quantify proteins by controlling methionine oxidation, improving reproducibility for disease research and diagnostics. The technique enhances peptide selection and mass spectrometry reliability.
Area of Science:
- Biochemistry
- Analytical Chemistry
- Proteomics
Background:
- Post-translational modifications (PTMs) are crucial for protein function but challenging to quantify accurately.
- Methionine oxidation during sample prep causes variability, excluding methionine-containing peptides from quantitative analysis.
- This limits coverage, especially for PTMs near methionine residues.
Purpose of the Study:
- To develop a robust strategy to eliminate methionine oxidation's impact on peptide quantification.
- To enable accurate measurement of methionine-containing peptides for broader quantitative coverage.
Main Methods:
- Developed a method to normalize sulfur element valence states by controlled oxidation of methionine residues.
- Optimized reaction parameters (H2O2 concentration, time, temperature) for complete and consistent methionine oxidation.
- Utilized two α-S2-casein peptides with phosphorylation sites and methionine as model analytes.
Main Results:
- Controlled oxidation stabilized methionine-containing peptides, eliminating variability.
- Achieved high precision with drastically reduced relative standard deviations (RSDs) from >15% to 0.24% and 1.42%.
- Accurately determined phosphorylation occupancies as 98.72% and 0.42%.
Conclusions:
- The developed method is straightforward, reproducible, and effective for quantifying methionine-containing peptides.
- Mitigates methionine-induced variability, expanding the range of quantifiable peptides.
- Enhances the reliability of protein metrology in mass spectrometry for research and diagnostics.
