Propagating Error through Traveling-Wave Ion Mobility Calibration

Area of Science:

• Biophysical Chemistry
• Structural Biology
• Mass Spectrometry

Background:

• Native mass spectrometry (MS) analyzes protein complexes in the gas phase, but ionization alters structures from solution.
• Gas-phase protein conformations offer insights into solution-phase energy landscapes.
• Ion mobility (IM) spectrometry characterizes gas-phase protein structures.

Purpose of the Study:

• To investigate gas-phase protein conformations using ion mobility spectrometry.
• To assess the impact of calibration uncertainty on traveling-wave ion mobility spectrometry (TWIMS) measurements.
• To improve the comparability of gas-phase protein structures derived from TWIMS with other experimental and computational data.

Main Methods:

• Calibrated a traveling-wave ion mobility spectrometry (TWIMS) instrument.
• Derived collision cross-section (CCS) values (TWCCS$_{N2}$ and TWCCS$_{N2→He}$) for four proteins.
• Propagated calibration error through TWIMS measurements.

Main Results:

• Reporting TWIMS collision cross-section (CCS) values with propagated calibration uncertainty significantly increased agreement with literature drift tube CCS (DT-CCS) values.
• This approach provides a more comprehensive assessment of protein ion gas-phase conformations.
• Four proteins (cytochrome c, ubiquitin, apo-myoglobin, holo-myoglobin) were analyzed.

Conclusions:

• Accurate error propagation in TWIMS calibration is crucial for reliable structural analysis.
• Improved gas-phase structural data enhances comparisons between native IM-MS and other structural datasets.
• This method refines the understanding of protein conformational landscapes in the gas phase.