Journal
JOURNAL OF PROTEOME RESEARCH
Volume 17, Issue 2, Pages 926-933Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acs.jproteome.7b00622
Keywords
electron capture dissociation; middle-down; top-down; bottom-up; Orbitrap; Q Exactive; disulfide; protein; mass spectrometry; post-translational modifications
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Funding
- large-scale proteomics facility Proteins@Work [184.032.201]
- Netherlands Organization for Scientific Research (NWO)
- European Union Horizon program FET-OPEN project MSmed [686547]
- Danish Agency for Science, Technology and Innovation
- Novo Nordisk STAR program
- Novo Nordisk Foundation [NNF14CC0001]
- NIH SBIR [R44 GM122131-01]
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As the application of mass spectrometry intensifies in scope and diversity, the need for advanced instrumentation addressing a wide variety of analytical needs also increases. To this end, many modern, top-end mass spectrometers are designed or modified to include a wider range of fragmentation technologies, for example, ECD, ETD, EThcD, and UVPD. Still, the majority of instrument platforms are limited to more conventional methods, such as CID and HCD. While these latter methods have performed well, the less conventional fragmentation methods have been shown to lead to increased information in many applications including middle-down proteomics, top-down proteomics, glycoproteomics, and disulfide bond mapping. We describe the modification of the popular Q Exactive Orbitrap mass spectrometer to extend its fragmentation capabilities to include ECD. We show that this modification allows >= 85% matched ion intensity to originate from ECD fragment ion types as well as provides high sequence coverage (>= 60%) of intact proteins and high fragment identification rates with similar to 70% of ion signals matched. Finally, the ECD implementation promotes selective disulfide bond dissociation, facilitating the identification of disulfide-linked peptide conjugates. Collectively, this modification extends the capabilities of the Q Exactive Orbitrap mass spectrometer to a range of new applications.
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