Development of novel isobaric tags enables accurate and sensitive multiplexed proteomics using complementary ions

Area of Science:

• Proteomics
• Analytical Chemistry
• Biochemistry

Background:

• High-throughput proteome analysis is crucial for understanding biological responses and diseases.
• Current isobaric labeling methods (e.g., TMT, iTRAQ) suffer from ratio distortion due to low-mass reporter ions and complex samples.
• Peptide coelution and cofragmentation in complex matrices limit quantification accuracy.

Purpose of the Study:

• To develop a novel isobaric labeling strategy for accurate high-throughput proteome quantification.
• To overcome the limitations of existing methods in complex biological samples.
• To enable sensitive and interference-free relative quantification.

Main Methods:

• Development of novel isobaric tags: dimethylated leucine complementary ion (DiLeuC).
• Utilizing high-mass complementary ions (remanent peptide segments) for quantification.
• Validation using a two-proteome model (yeast spiked into human proteome) and single-cell proteome analysis.

Main Results:

• DiLeuC tags provide interference-free relative quantification using precursor-specific, high-mass complementary ions.
• Demonstrated high quantification accuracy in a complex two-proteome model.
• Successfully applied the strategy to sensitive single-cell proteome analysis.

Conclusions:

• The DiLeuC method offers a significant advancement for accurate and sensitive proteome quantification.
• This novel approach overcomes major limitations of current isobaric labeling techniques.
• DiLeuC has broad potential applications in various fields of biological research, including single-cell analysis.