Peptide and protein sequence analysis by electron transfer dissociation mass spectrometry

Area of Science:

• Analytical Chemistry
• Biochemistry
• Mass Spectrometry

Background:

• Tandem mass spectrometry (MS/MS) is crucial for peptide sequencing.
• Electron capture dissociation (ECD) is an effective MS/MS technique for peptide fragmentation.
• Developing alternative fragmentation methods for MS/MS is an ongoing area of research.

Purpose of the Study:

• To demonstrate peptide sequence analysis using gas-phase ion/ion chemistry combined with MS/MS.
• To present modifications to a radio frequency quadrupole linear ion trap (QLT) for enabling ion/ion chemistry.
• To describe automated acquisition of electron transfer dissociation (ETD) MS/MS spectra for phosphopeptides.

Main Methods:

• Utilizing singly charged anthracene anions for electron transfer to multiply protonated peptides.
• Employing a modified radio frequency quadrupole linear ion trap (QLT) for ion/ion reactions.
• Coupling nanoflow high-performance liquid chromatography (HPLC) with automated MS/MS acquisition.

Main Results:

• Peptide backbone fragmentation was induced via electron transfer, analogous to ECD pathways.
• High-quality, single-scan ETD MS/MS spectra were obtained for phosphopeptides.
• The modified QLT successfully facilitated the ion/ion chemistry for peptide analysis.

Conclusions:

• Gas-phase ion/ion chemistry in conjunction with MS/MS provides a viable method for peptide sequencing.
• The described modifications and automated acquisition enable efficient analysis of challenging peptide samples, including phosphopeptides.
• Electron transfer dissociation represents a powerful complementary technique to existing MS/MS methods.