Trapped Ion Mobility Spectrometry of Native Macromolecular Assemblies

Area of Science:

• Analytical Chemistry
• Biophysics
• Structural Biology

Background:

• Native mass spectrometry (MS) and ion mobility spectrometry-MS (IMS-MS) are crucial for studying macromolecular assemblies.
• Understanding the biochemical and biophysical functions of these assemblies requires detailed structural information.

Purpose of the Study:

• To introduce a new generation trapped ion mobility spectrometer (TIMS) designed for enhanced trapping of high-molecular-weight (MW) macromolecular assemblies.
• To demonstrate the capabilities of this novel TIMS analyzer for structural elucidation.

Main Methods:

• Development of a compact 4 cm TIMS analyzer featuring a convex electrode and quadrupolar geometry.
• Extended mobility range (K0 = 0.185–1.84 cm2·V−1·s−1) enabling trapping of high-MW species in native states (lower charge states).
• Utilized variable scan rate mobility measurements, high-mobility resolution, and ion-neutral collision cross-section (CCS N) measurements.

Main Results:

• The novel TIMS successfully trapped a wide range of standards, including CsI clusters, proteins, antibodies, protein-DNA complexes, and macromolecular assemblies (GroEL, RNA polymerase).
• Demonstrated effective trapping of species with masses up to m/z 19,000 and CCS up to 22,000 Å2.
• Achieved high precision with <0.6% relative standard deviation (RSD) in CCS measurements.

Conclusions:

• The new convex electrode TIMS geometry significantly extends trapping capabilities for high-MW macromolecular assemblies.
• This instrument offers improved structural elucidation of complex biological systems through enhanced mass spectrometry analysis.
• The TIMS demonstrates ease of operation and broad applicability across diverse biomolecular assemblies.