How to Deal With Internal Fragment Ions?

Area of Science:

• Proteomics
• Analytical Chemistry
• Computational Biology

Background:

• Tandem mass spectrometry (MS/MS) generates fragment ions for peptide and protein analysis.
• N- and C-terminal ions are routinely identified, but internal fragment ions are challenging to annotate.
• Internal fragments are vital for long peptides and proteoforms, offering insights into sequence and modifications.

Purpose of the Study:

• To analyze the contribution of internal fragment ions in middle-down and top-down mass spectrometry.
• To introduce a novel graph-based approach for annotating complex internal fragment ions.
• To improve the accuracy and coverage of proteoform sequencing and post-translational modification localization.

Main Methods:

• Analysis of internal fragment ion contributions in middle-down and top-down mass spectra.
• Development of a graph-based representation for comparing multiple candidate proteoforms.
• Assessment of candidate annotations within fragment ion spectra using the graph model.

Main Results:

• Internal fragment ions significantly enhance amino acid sequence coverage for polypeptides and proteins.
• The graph-based method effectively manages the complexity of internal fragments, reducing false positives.
• Accurate localization of post-translational modifications is improved using internal fragment ion data.

Conclusions:

• Internal fragment ions provide valuable information for complex proteoform characterization.
• The graph-based annotation approach offers a general solution for processing complex tandem mass spectra.
• This method enhances proteoform sequencing and the characterization of post-translational modifications by mass spectrometry.