Proteoform Differentiation using Tandem Trapped Ion Mobility, Electron Capture Dissociation, and ToF Mass Spectrometry

Kevin Jeanne Dit Fouque, Desmond Kaplan, Valery G. Voinov, Frederik H.V. Holck, Ole N. Jensen, Francisco Fernandez-Lima*

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Comprehensive characterization of post-translationally modified histone proteoforms is challenging due to their high isobaric and isomeric content. Trapped ion mobility spectrometry (TIMS), implemented on a quadrupole/time-of-flight (Q-ToF) mass spectrometer, has shown great promise in discriminating isomeric complete histone tails. The absence of electron activated dissociation (ExD) in the current platform prevents the comprehensive characterization of unknown histone proteoforms. In the present work, we report for the first time the use of an electromagnetostatic (EMS) cell devised for nonergodic dissociation based on electron capture dissociation (ECD), implemented within a nESI-TIMS-Q-ToF mass spectrometer for the characterization of acetylated (AcK18 and AcK27) and trimethylated (TriMetK4, TriMetK9 and TriMetK27) complete histone tails. The integration of the EMS cell in a TIMS-q-TOF MS permitted fast mobility-selected top-down ECD fragmentation with near 10% efficiency overall. The potential of this coupling was illustrated using isobaric (AcK18/TriMetK4) and isomeric (AcK18/AcK27 and TriMetK4/TriMetK9) binary H3 histone tail mixtures, and the H3.1 TriMetK27 histone tail structural diversity (e.g., three IMS bands atz= 7+). The binary isobaric and isomeric mixtures can be separated in the mobility domain withR IMS> 100 and the nonergodic ECD fragmentation permitted the PTM localization (sequence coverage of ∼86%). Differences in the ECD patterns per mobility band of thez= 7+ H3 TriMetK27 molecular ions suggested that the charge location is responsible for the structural differences observed in the mobility domain. This coupling further enhances the structural toolbox with fast, high resolution mobility separations in tandem with nonergodic fragmentation for effective proteoform differentiation.

TidsskriftAnalytical chemistry
Udgave nummer27
Sider (fra-til)9575–9582
StatusUdgivet - 13. jul. 2021

Bibliografisk note

Funding Information:
The authors at FIU acknowledge the financial support from the National Science Foundation Division of Chemistry, under CAREER award CHE-1654274, with cofunding from the Division of Molecular and Cellular Biosciences to FFL and funding from National Institutes of General Medicine (R01GM134247). We thank Dr. Mark E. Ridgeway and Dr. Melvin A. Park for helpful discussions during the TIMS-q-EMS-ToF MS development.

Publisher Copyright:
© 2021 American Chemical Society.


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