Intact Mass Proteomics Using a Proteoform Atlas

Area of Science:

• Proteomics
• Mass Spectrometry
• Molecular Biology

Background:

• Top-down proteomics characterizes intact proteoforms using tandem mass spectrometry (MS/MS).
• This method provides detailed molecular information but is limited by slow identification speeds due to reliance on MS/MS.
• Faster proteoform identification is crucial for analyzing complex biological systems.

Purpose of the Study:

• To develop and validate a faster proteoform identification strategy using a system-specific Proteoform Atlas.
• To enable rapid MS1-only identifications of proteoforms in E. coli.
• To assess the efficiency of the Atlas-based approach compared to traditional top-down proteomics.

Main Methods:

• Constructed an E. coli ribosomal Proteoform Atlas via deep top-down mass spectrometry analysis.
• The Atlas contains 2099 proteoforms from 52 E. coli ribosomal proteins.
• Applied the Atlas to identify proteoforms in E. coli exposed to cold stress using MS1-only scans.

Main Results:

• Successfully built a comprehensive E. coli ribosomal Proteoform Atlas.
• Demonstrated confident MS1-only identification of proteoforms in cold-stressed E. coli using the Atlas.
• Achieved up to 77% increased identification speed compared to traditional top-down MS methods.

Conclusions:

• A Proteoform Atlas enables significantly faster proteoform identification in E. coli.
• The MS1-only identification strategy is efficient and reliable for systems with established Atlases.
• This approach accelerates proteomic analysis, particularly in response to biological perturbations.