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Updated: Jul 27, 2025
07:33
Analyzing Protein Architectures and Protein-Ligand Complexes by Integrative Structural Mass Spectrometry
Published on: October 15, 2018
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Orbitrap-Based Mass and Charge Analysis of Single Molecules
Evolène Deslignière1,2, Amber Rolland1,2, Eduard H T M Ebberink1,2
1Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Accounts of Chemical Research
|June 6, 2023
Summary
Native mass spectrometry struggles with complex protein mixtures. Orbitrap-based charge detection mass spectrometry (CDMS) now analyzes single molecules, overcoming limitations for heterogeneous assemblies and enhancing protein mass analysis sensitivity and resolution.
Area of Science:
- Biochemistry
- Analytical Chemistry
- Mass Spectrometry
Background:
- Native mass spectrometry is crucial for analyzing intact proteins and biomolecular assemblies.
- Traditional methods face challenges with heterogeneous complexes due to stoichiometry, subcomplexes, and post-translational modifications.
- Current techniques require millions of molecules, limiting sensitivity.
Purpose of the Study:
- To introduce and detail the advancements in Orbitrap-based charge detection mass spectrometry (CDMS) for analyzing complex protein assemblies.
- To overcome the limitations of traditional mass spectrometry in handling heterogeneous samples and low molecule counts.
- To demonstrate the capabilities of Orbitrap-based CDMS in various biological applications.
Main Methods:
- Development and optimization of Orbitrap-based mass analyzers with extended mass range (EMR).
- Implementation of single-molecule detection principles for charge detection mass spectrometry (CDMS).
- Application of Orbitrap-based CDMS to analyze heterogeneous protein complexes, including glycoprotein assemblies and viral vectors.
Main Results:
- Orbitrap-based CDMS enables single-ion measurements, providing sufficient signal for mass analysis.
- The technique circumvents traditional charge state inferencing, allowing analysis of highly heterogeneous protein assemblies.
- Demonstrated applications include assessing viral vector cargo, immune complex formation, and SARS-CoV-2 spike protein mass.
Conclusions:
- Orbitrap-based CDMS significantly enhances the analysis of complex and heterogeneous protein assemblies.
- Single-molecule detection offers unprecedented insights into ion behavior and stability within the mass analyzer.
- The goal is to make Orbitrap-based CDMS a mainstream technique, further improving sensitivity and mass resolving power.
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