Effective discrimination of gas-phase peptide conformers using TIMS-ECD-ToF MS/MS

Area of Science:

• Analytical Chemistry
• Biochemistry
• Structural Biology

Background:

• Peptide structure and conformation are crucial for biological function.
• Ion mobility and mass spectrometry fragmentation techniques offer insights into molecular structure.
• Correlating these techniques can provide a more comprehensive understanding of peptide conformations.

Purpose of the Study:

• To correlate structural information from ion mobility and electron capture dissociation (ECD)/collision-induced dissociation (CID) fragmentation.
• To investigate the structural heterogeneity and conformational dynamics of model peptides.
• To evaluate the utility of a TIMS-q-EMS-ToF MS/MS platform for detailed peptide structural analysis.

Main Methods:

• Utilized a novel TIMS-q-EMS-ToF MS/MS platform.
• Employed high ion mobility resolving power (R ~115-145) to resolve peptide conformers.
• Analyzed fast online ECD fragmentation patterns specific to each ion mobility band.

Main Results:

• Observed structural heterogeneity in model peptides (substance P, bradykinin, angiotensin I, AT-Hook 3) via multi-component ion mobility profiles.
• Identified conformers likely involving cis/trans isomerizations at X-Pro peptide bonds.
• Demonstrated comparable ECD fragmentation efficiencies between ion mobility-selected and traditional FT-ICR ECD MS/MS, with differences in radical fragment ratios attributed to detection time.

Conclusions:

• The TIMS-q-EMS-ToF MS/MS platform provides complementary structural information on intramolecular interactions stabilizing gas-phase peptide conformations.
• Online coupling of ion mobility with ECD fragmentation enhances structural characterization beyond individual techniques.
• This integrated approach offers deeper insights into peptide structural dynamics and isomerism.