Protease- and Acid-catalyzed Labeling Workflows Employing 18O-enriched Water

Overview

This presentation demonstrates workflows for stable isotope labeling of protein digests using 18O-enriched water. The methods include protease-assisted and acid-catalyzed workflows, which facilitate the incorporation of 18O atoms for proteomic studies.

Key Study Components

Area of Science

• Proteomics
• Stable Isotope Labeling
• Mass Spectrometry

Background

• Stable isotope labeling is crucial for quantitative and qualitative proteomics.
• 18O-enriched water is used to introduce stable isotopes into proteins.
• Protease-assisted workflows utilize proteolytic cleavage for isotope incorporation.
• Acid-catalyzed workflows allow for incorporation at low pH.

Purpose of Study

• To demonstrate versatile workflows for stable isotope labeling.
• To enhance quantitative and qualitative analysis in proteomics.
• To showcase the incorporation of 18O atoms in protein studies.

Methods Used

• Protease-assisted workflows with 50% 18O-enriched water.
• Acid-catalyzed workflows for functional group labeling.
• Mass spectrometric data acquisition for spectral analysis.
• Time course experiments to track peptide cleavage and labeling.

Main Results

• Successful incorporation of 18O atoms into peptide species.
• Generation of spectral time plots for peptide cleavage reactions.
• Identification of labeled cleavage products using mass spectrometry.
• Demonstration of the versatility of the workflows in proteomics.

Conclusions

• The workflows provide effective tools for proteomic studies.
• 18O labeling enhances the understanding of protein dynamics.
• These methods can be applied to various proteomic research areas.

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