Gas-phase bovine ubiquitin cation conformations resolved by gas-phase hydrogen/deuterium exchange rate and extent

Michael A. Freitas, Christopher L. Hendrickson, Mark R. Emmett, Alan G. Marshall

Research output: Contribution to journalArticlepeer-review

138 Scopus citations


The gas-phase hydrogen/deuterium exchange of [M + nH]n+ (n = 5-13) ions of bovine ubiquitin with the H/D exchange reagent D2O are examined by electrospray ionization Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry. All of the odd or all of the even charge states were isolated by stored waveform inverse Fourier transform excitation and simultaneously reacted with D2O leaked steadily into the ICR cell for reaction periods ranging from 1 s to 1 h. Different gas-phase protein conformations could be resolved according to difference in extent of H/D exchange. The 5+ and 6+ charge states display broad distributions of conformations ranging from 0-80% deuterium incorporation. In contrast, each of the higher charge states, 7-11+ and 13+, displays a single major isotopic distribution, whereas the 12+ charge state separates into two isotopic distributions of comparable abundance. In general, H/D exchange rates decrease with increasing charge state. External electrospray ionization source conditions (capillary current and external accumulation period) were varied while observing the conformational distribution of the 7+ charge state: increased heating in either region reduced the number of slow-exchanging conformations. At 9.4 T, it is possible to trap a large number of ions for a long reaction period (up to 1 h) at relatively high pressure (2 × 10-7 Torr). These results demonstrate the capability of FT-ICR mass analysis following gaseous H/D exchange of electrosprayed proteins to disperse different gas-phase protein conformations for subsequent isolation and characterization.

Original languageEnglish (US)
Pages (from-to)565-575
Number of pages11
JournalInternational Journal of Mass Spectrometry
StatePublished - 1999
Externally publishedYes


  • Fourier transform
  • Gas phase ion-molecule reactions
  • Hydrogen-deuterium exchange
  • Ion cyclotron resonance
  • Mass spectrometry
  • Protein

ASJC Scopus subject areas

  • Instrumentation
  • Condensed Matter Physics
  • Spectroscopy
  • Physical and Theoretical Chemistry


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