A Multiplexed SEC-MS Approach to Systematically Study the Interplay Between Protein Assembly-States and...

Area of Science:

• Molecular Biology
• Proteomics
• Biochemistry

Background:

• Protein molecular interactions and post-translational modifications (PTMs) are co-dependent and co-regulate biological processes.
• A systematic method to simultaneously study PTMs and protein assembly states from the same sample is lacking.

Purpose of the Study:

• To introduce SEC-MX (Size Exclusion Chromatography fractions MultipleXed), a novel global quantitative method.
• To enable simultaneous characterization of PTMs and assembly states.
• To facilitate quantitative differential comparisons between biological conditions.

Main Methods:

• Combining Size Exclusion Chromatography (SEC) with PTM-enrichment techniques.
• Global quantitative analysis of phosphopeptides and their associated assembly states.
• Application to HEK293 and HCT116 cell lines for proof-of-concept.

Main Results:

• Generated a comprehensive dataset mapping thousands of phosphopeptides and their assembly states.
• Revealed intricate relationships between phosphorylation events and protein assembly states.
• Provided testable hypotheses for further investigation into protein function and regulation.

Conclusions:

• SEC-MX is established as a valuable tool for studying protein function and regulation.
• The method goes beyond traditional abundance changes to explore PTMs and assembly states.
• Enables deeper insights into the interplay between protein modifications and molecular interactions.