Increased Single-Spectrum Top-Down Protein Sequence Coverage in Trapping Mass Spectrometers with Chimeric Ion Loading

Area of Science:

• Analytical Chemistry
• Biochemistry
• Mass Spectrometry

Background:

• Fourier transform mass spectrometry (FT-MS) offers high resolution and accuracy.
• Top-down proteomics requires efficient methods for comprehensive protein analysis.

Purpose of the Study:

• To introduce and validate a novel ion loading technique called "chimeric ion loading" for FT-ICR-MS.
• To improve protein sequence coverage and acquisition efficiency in top-down proteomics.

Main Methods:

• Utilized a 21 T FT-ICR mass spectrometer.
• Applied multiple dissociation techniques (collision-induced dissociation [CID] and electron-transfer dissociation [ETD]) and multiple precursor ions within a single transient acquisition.
• Analyzed MCF7 cell lysate using liquid chromatography-mass spectrometry (LC-MS).

Main Results:

• Chimeric ion loading significantly improved mean sequence coverage (from 15% with CID-only to 33% with chimeric approach).
• The technique demonstrated multiplexing capabilities for multiple charge states and reaction periods.
• Acquisition time was reduced and sample consumption decreased.

Conclusions:

• Chimeric ion loading is an effective strategy for enhancing top-down proteomics efficiency and data quality.
• This method holds potential for broader applications in tandem mass spectrometry beyond proteomics.