Supercritical fluid chromatography reduction of hydrogen/deuterium back exchange in solution-phase hydrogen/deuterium exchange with mass spectrometric analysis

Mark R. Emmett, Sasa Kazazic, Alan G. Marshall, Wei Chen, Stone D.H. Shi, Ben Bolaños, Michael J. Greig

Research output: Contribution to journalArticlepeer-review

26 Scopus citations

Abstract

The single biggest problem with solution-phase H/D exchange as a mass spectrometric probe of surface exposure in a protein (or protein complex) is back-exchange of H for D after the initial H/D exchange has been quenched. Back-exchange results in loss of pertinent data and also greatly hampers data analysis. Previously, very fast, cold (0-4 °C) HPLC was performed to help reduce back-exchange, but calculated back-exchange still averages ∼30%. In this report, supercritical fluid chromatography replaces HPLC as the desalting/separation technique prior to mass analysis, providing a dramatic reduction in back-exchange compared to the fast, cold HPLC methods.

Original languageEnglish (US)
Pages (from-to)7058-7060
Number of pages3
JournalAnalytical Chemistry
Volume78
Issue number19
DOIs
StatePublished - Oct 1 2006
Externally publishedYes

ASJC Scopus subject areas

  • Analytical Chemistry

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