Mass Defect-Based Dimethyl Pyrimidinyl Ornithine (DiPyrO) Tags for Multiplex Quantitative Proteomics

Area of Science:

• Proteomics
• Mass Spectrometry
• Chemical Biology

Background:

• Quantitative proteomics is crucial for biological research.
• Existing methods for multiplexed peptide quantification can be limited by mass spectral complexity and resolving power requirements.

Purpose of the Study:

• To develop a novel, compact, and versatile chemical tag for highly multiplexed peptide quantification.
• To enable quantitative proteomics at the MS¹ level with reduced reliance on high resolving power.

Main Methods:

• Design and synthesis of the dimethyl pyrimidinyl ornithine (DiPyrO) tag with isotopic configurations.
• Characterization of DiPyrO-labeled proteome analysis using nanoLC-MS².
• Demonstration of duplex and triplex quantitative proteomics using high-resolution mass spectrometry (Orbitrap Elite and Fusion Lumos).

Main Results:

• The DiPyrO tag imparts a tunable mass defect (5.8–45.3 mDa per tag) between labeled peptides.
• Lysine-containing peptides labeled with two DiPyrO tags achieve up to 90.6 mDa mass defect, reducing resolving power needs.
• Successful duplex and triplex quantitative proteomics demonstrated on Orbitrap platforms.

Conclusions:

• DiPyrO is a novel chemical tag enabling highly multiplexed peptide quantification at the MS¹ level.
• The tag's design allows for confident quantitative analyses on various LTQ-Orbitrap platforms with reduced resolving power.
• DiPyrO offers a versatile tool for advancing quantitative proteomics research.