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Top-down protein identification using a time-of-flight mass spectrometer and data independent acquisition.
- Source :
-
International Journal of Mass Spectrometry . Jan2019, Vol. 435, p136-144. 9p. - Publication Year :
- 2019
-
Abstract
- Highlights • Top-down proteomics with an LC-TOF analyzer is demonstrated. • Data-independent acquisition (DIA) for top-down MS is shown. • All subunits and several PTMs for the h20S proteasome are identified. Abstract Top-down mass spectrometry and direct dissociation of gas phase intact proteins have been demonstrated to be a powerful platform for identifying proteins from complex mixtures and for elucidating post-translational modifications (PTMs). Fragmentation of proteins in the atmospheric pressure/vacuum interface of the electrospray ionization mass spectrometer is an effective dissociation technique that can be utilized for on-line HPLC top-down analysis. We demonstrate the capability to perform intact protein identifications in a single-stage time-of-flight (TOF) mass spectrometer in a data independent (DIA) acquisition fashion by rapidly switching the in-source dissociation (ISD) energy during protein elution from a liquid chromatography (LC) column. The intact protein and product ion masses obtained at low and high ISD energies, respectively, were measured using a TOF mass analyzer. By coupling on-line protein separations to dissociation in the atmospheric pressure/vacuum interface region of the mass spectrometer, we identified proteins in simple complexity mixtures, including subunits from the human 20S proteasome complex, and PTMs such as phosphorylation and N-terminal acetylation events. This proof-of-principle study demonstrates that a data-independent pseudo-MS/MS method could be a relatively inexpensive platform for top-down MS. [ABSTRACT FROM AUTHOR]
- Subjects :
- *PROTEOMICS
*PROTEASOMES
*GAS phase reactions
*PROTEINS
*LIQUID chromatography
Subjects
Details
- Language :
- English
- ISSN :
- 13873806
- Volume :
- 435
- Database :
- Academic Search Index
- Journal :
- International Journal of Mass Spectrometry
- Publication Type :
- Academic Journal
- Accession number :
- 133366874
- Full Text :
- https://doi.org/10.1016/j.ijms.2018.10.023