Combining Targeted and Untargeted Data Acquisition to Enhance Quantitative Plant Proteomics Experiments

Area of Science:

• Proteomics
• Mass Spectrometry
• Biochemistry

Background:

• Quantitative proteomics typically involves either targeted analysis of specific proteins or broad, untargeted profiling.
• Previous work indicated that integrating targeted and untargeted approaches offers significant advantages.

Purpose of the Study:

• To describe the creation and implementation of combined targeted and untargeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods.
• To demonstrate the benefits of a targeted data acquisition/data-dependent acquisition (TDA/DDA) strategy for quantitative proteomics.

Main Methods:

• Development of TDA/DDA LC-MS/MS methods on standard mass spectrometry platforms.
• Generation of inclusion lists for targeted protein quantification using free software.
• Application of TDA/DDA to a model plant proteomics experiment.

Main Results:

• TDA/DDA improved the quantification of both targeted and untargeted proteins compared to conventional data-dependent acquisition.
• Increased reproducibility in untargeted protein quantification without reducing the total number of proteins identified.
• Successful application in a plant proteomics study.

Conclusions:

• The TDA/DDA strategy offers a valuable alternative to conventional data-dependent acquisition for quantitative proteomics.
• This method enhances the reliability of protein quantification and is compatible with various labeling techniques.
• TDA/DDA can facilitate the creation of assay libraries for data-independent acquisition analysis.