Retention Time Alignment for Protein Turnover Studies Using Heavy Water Metabolic Labeling

Area of Science:

• Proteomics
• Analytical Chemistry
• Biochemistry

Background:

• Retention time (RT) alignment is crucial for accurate protein identification and quantification in mass spectrometry-based proteomics.
• Matched between runs (MBR) is an approach that leverages RT alignment to transfer peptide identifications across experiments, enhancing data completeness.
• RT alignment has not been widely applied to quantitative proteomics studies involving stable isotope labeling, such as metabolic labeling with deuterium oxide (D2O).

Purpose of the Study:

• To develop and evaluate an approach integrating RT alignment with peptide quantification for proteome turnover studies using D2O metabolic labeling.
• To improve the reliability and coverage of peptide identification and quantification in D2O-labeled proteomic experiments.

Main Methods:

• Utilized correlation-optimized time warping for RT alignment of liquid chromatography-mass spectrometry (LC-MS) data.
• Integrated RT alignment with the matched between runs (MBR) strategy for peptide transfer and quantification.
• Applied the approach to analyze proteome turnover in samples labeled with deuterium oxide (D2O).

Main Results:

• The integrated RT alignment and MBR approach significantly improved labeling time point coverage.
• The method demonstrated particular effectiveness for experiments with long labeling durations.
• Enhanced the ability to identify and quantify peptides in D2O-labeled samples, addressing challenges posed by altered isotope profiles.

Conclusions:

• Incorporating RT alignment with MBR is a valuable strategy for robust proteome turnover analysis using D2O metabolic labeling.
• This approach enhances the efficiency and comprehensiveness of quantitative proteomics, particularly for dynamic biological processes.
• The developed method facilitates more accurate insights into protein dynamics and metabolic pathways.